Study design
This was a partly randomized, parallel, single-center study comprising patients with AF scheduled for mitral valve surgery. A group of patients ineligible for AF ablation were included as controls and were not subject to randomization.
Study population
Of 63 consecutive patients, 3 declined participation. Sixty patients with preoperative AF scheduled for mitral valve surgery were enrolled consecutively between Sept 2013 and October 2015 at Linkoping University Hospital. Patients judged to benefit from AF ablation were randomly assigned, in a 1:1 ratio in blocks of 10, to undergo concomitant AF ablation with either a cryodevice (cryo group, n = 20) or RF device (RF group, n = 21). Patients with no AF symptoms, and those for whom additional aortic cross clamp time and thereby prolonged ischemia was regarded as too hazardous, underwent mitral valve surgery without AF ablation (no-maze group, n = 19). The decision to ablate or not was made before enrollment by a group of experts in arrhythmia, consisting of cardiologists and cardiac surgeons. The study was approved by the Regional Ethical Review Board (2012/371 − 31) and patients were recruited after written, informed consent by the surgeon the day before surgery. This study is the first part of the registered clinical trial: DOI 10.1186/ISRCTN14454361.
Ablation methods
For cryoablation, the argon powered Cardioblate CryoFlex Surgical Ablation Probe (Medtronic Inc., Minneapolis, MN) was used. The probe was applied for 120 s and the temperature fell rapidly (Joule-Thompson effect) to between − 130 °C and − 150 °C at each ablation line. The left atrial lesions consisted of three lines for the left atrium and three for the right atrial wall and followed the Cox IV pattern [16–18]. All but the intercaval lesions were created during cardioplegic arrest. For RF ablation, the Cardioblate BP2 Irrigated Bipolar Surgical Ablation System (Medtronic Inc., Minneapolis, MN) was used. This system uses irrigation and impedance-based power adjustments to reach tissue temperatures between 50 °C and 80 °C during ablation, according to the manufacturer. Each line was subject to at least three complete ablation periods. During RF ablation, the right and the left pulmonary vein orifices were isolated pairwise and epicardially on bypass [19], but before cardioplegia was given. During cardioplegic arrest, the left atrial ablation was completed according to the Cox IV pattern [20].
Procedures
All patients were operated on by the same surgeon. Myocardial arrest was induced and maintained with combined antegrade and retrograde cold blood cardioplegia. Biatrial lesions were induced, during cardioplegia arrest, in all ablation patients except for one in the cryo group and three in the RF group, who underwent only left-sided ablations. The left atrial appendage was stapled and excised with an Endo GIA Reinforced device (Medtronic Inc., Minneapolis, MN). The surgical procedures and patient characteristics are summarized in Table 1.
Blood sampling and biochemical analyses
Blood was collected preoperatively on the day before surgery (Pre), at the end of surgery (T0), 3 h (T3), 20 h (T20), and 3 days (D3) postoperatively. Troponin T (Tn-T) was analyzed at Pre, T0, T3, T20, and D3. Creatine kinase isoenzyme MB (CK-MB) and aspartate aminotransferase (ASAT) were analyzed at Pre, T0, T3, and T20. C-reactive protein (CRP) and alanine aminotransferase (ALAT) were analyzed at Pre and T20. NT-proBNP was analyzed at Pre, T20, and D3.
CK-MB, Tn-T, and NT-proBNP was analyzed in serum samples using an electrochemiluminescence immunoassay. A particle-enhanced immunoturbidimetric assay was used for CRP measurement in plasma samples preoperatively and postoperatively, and pyridoxal phosphate activation for ASAT and ALAT. The cobas c701 and e602 analyzers (Roche Diagnostics GmbH, Mannheim, Germany) were used. Additional biomarkers were analyzed using the Proseek Multiplex CVD II panel (Olink Proteomics, Uppsala, Sweden), simultaneously measuring 92 proteins. This analysis provides very high sensitivity and specificity as it is performed using a matched pair of antibodies coupled to unique oligonucleotides and measured by quantitative real-time PCR. For this study, blood samples from Pre, T0, T3, and T20 were fractionated and plasma was stored at − 70 °C before being sent to the Clinical Biomarkers National Facility (Science for Life Laboratory, Uppsala) for analysis.
In this survey, interleukins 6 and 18 (IL-6 and IL-18) were selected as markers for inflammation, and serine/threonine-protein kinase 4 (STK4) also known as MST1, proteinase activated receptor 1 (PAR-1), heat shock 27 kDa protein (HSP27), and TNF-related apoptosis-inducing ligand receptor 2 (TRAIL-R2) were selected as markers of cellular stress/apoptosis. Data from the Proseek Multiplex CVD II panel are presented as the differences in normalized protein expression (NPX) between preoperative levels and T0, T3, and T20 (ΔT0, ΔT3, and ΔT20). The log-2 transformed data were linearized before calculating differences from preoperative values. Data from 3.8% of the samples were excluded from analysis due to assay failure or failure to pass quality control.
Statistical methods
Sample-size calculation was based on the difference in markers for myocardial injury in pilot patients with and without concomitant cryoablation. The number of patients needed in each group to detect such a large difference was 3. We included 60 patients in the study to ensure at least 17 patients each in the cryoablation and RF ablation groups, enough to detect an effect size of 1. Continuous variables are presented as medians [25th to 75th percentile] and categorical variables as counts (percentages). For comparisons of the three groups, the Kruskal-Wallis test was used, taking into account multiple comparisons of mean ranks for all groups’ P values, and post-hoc comparisons of mean ranks of all pairs of groups when the Kruskal-Wallis test showed significance. The Spearman rank test was used for correlations between two variables. Multivariable linear regression models were used to find the factors influencing myocardial injury. The Tn-T and CK-MB at 3 h postoperatively and ASAT at 20 h postoperatively were chosen as dependent factors and log-10 transformed to align with the assumption of Gaussian distribution. Study groups, age, gender, cross clamp time (CCT), coronary artery disease, and the extent of ablation were tested as predictors. The models were fit using an enter method. Multicollinearity was found between the study groups and the extent of ablation. Therefore, the extent of ablation was excluded from the models. For comparison of the markers of myocardial injury between the three study groups over time-repeated measurements, analysis of variance was used. A P value < 0.05 was considered statistically significant. Statistical analyses were conducted with Statistica 10.0 (StatSoft Inc., Tulsa, OK) and SPSS 22.0 (SPSS, Inc., Chicago, IL).