Role of Preexisting Proarrhythmic Atrial Remodeling in Post-Coronary Artery Bypass Grafting Atrial Fibrillation

Consecutive patients with coronary artery disease hospitalized for an elective CABG procedure at the Emergency Institute for Cardiovascular Diseases and Transplantation Targu Mures between March 2016 and June 2017 were prospectively evaluated. Patients were included in the study if they were admitted to hospital for a first-time elective CABG procedure, if they were hemodynamically stable at the moment of inclusion, and if they were ≥18 years of age. Exclusion criteria were: concomitant valve replacement or other intervention planned to be performed within the same procedure, systemic inflammatory diseases, and treatment with steroids or other anti-inflammatory drugs. Patients with a history of AF were also excluded. Median sternotomy and standard extracorporeal circulation (ECC) were used in all patients. All patients included in the study signed written informed consent. The research protocol complied with the Helsinki Declaration and was approved by the local Ethics Committees.

Patients were divided into two groups (i.e., AF and no-AF), according to the presence or absence of post-CABG AF. For patients to be allocated to their according groups, all patients underwent continuous 12-lead Holter ECG monitoring from day-1 post-CABG up to the day of discharge using CardioMem 3000^® systems (GETEMED/GE Healthcare). Post-CABG AF was defined as the occurrence of AF episode(s) lasting ≥30 s during post-CABG hospitalization. Clinical characteristics (i.e., age, gender, body mass index [BMI], creatinine clearance), comorbidities (i.e., obesity, arterial hypertension, diabetes mellitus, history of stroke), and ongoing drug therapy were recorded for each patient. The CHA₂DS₂VASc and SYNergy between PCI with TAXUS™ and Cardiac Surgery (SYNTAX I) scores were also calculated. The duration of ECC, of aortic clamping, and the total duration of the CABG procedure were also recorded for each patient.

To assess the presence and severity of preexisting atrial remodeling and its relationship with post-CABG AF, all patients underwent (Figure 1) surface ECG recording and echocardiographic assessment on admission. Continuous 24-h 12-lead ECG Holter monitoring (CardioMem 3000^®; GETEMED/GE Healthcare) was also performed in all patients prior to CABG to evaluate the presence and severity of preexisting autonomic remodeling, as well as the presence and burden of pre-CABG atrial arrhythmic events. During the CABG procedure, the right atrial appendage, normally removed during fixation of the ECC cavo-atrial cannula and routinely considered ‘surgical waste’, was collected and divided into three samples. One sample was used for electrophysiological studies, one for histological analyses, and one for transcriptome analyses, as described below. To evaluate the occurrence of post-CABG AF, all patients underwent continuous post-CABG 12-lead ECG Holter monitoring, as described above. Clinically relevant postoperative inhospital outcomes were also recorded, as described below.

Figure 1

To assess the presence and severity of preexisting atrial electrical remodeling, P wave characteristics (i.e., duration, amplitude, fragmentation) were evaluated in lead II and Lewis leads using the surface ECG recordings obtained on admission. The burden of pre-CABG atrial arrhythmic events (i.e., atrial premature beats) was also evaluated using the continuous 24-h 12-lead ECG recordings obtained prior to CABG.

Electrophysiological study of a right atrial appendage sample was also performed. Upon collection, the sample was immediately placed into Krebs-Henseleit solution (118 mM NaCl, 4.7 mM KCl, 25 mM NaHCO₃, 2.5 mM CaCl₂, 1.2 mM MgSO₄, 1.2 mM KH₂PO₄, and 11 mM glucose) and transferred into the Steiert organ bath. After a 30-min equilibration period, the right atrial appendage sample was electrically stimulated and atrial action potentials were recorded. The amplitudes of the resting and of the action potential and the duration of depolarization and that of the action potential to 25% (APD25), 50% (APD50), and 90% (APD90) of complete repolarization were measured.

To evaluate a potential preexisting atrial electrical remodeling at a molecular level, the right atrial appendage sample collected for transcriptome analysis was rapidly immersed into RNA stabilizing solution (RNAlater; Thermo Fisher Scientific, Waltham, MA) and the mRNA levels of several genes previously associated with AF occurrence (i.e. CACNA1C, GJA5, KCNE2, KCNJ2, KCNQ1, and SCN5A) were analyzed. Total iPrep PureLink RNA Kits and iPrep Purification Tool (Thermo Fisher Scientific) were used to isolate RNA and the SuperScript VILO cDNA synthesis kit (Thermo Fisher Scientific) was used to perform reverse transcription. The mRNA expression levels of the six target genes were analyzed using TaqMan Gene Expression Assays (Thermo Fisher Scientific). All experiments were performed on a 7500 Fast Dx Real-Time PCR System (Applied Biosystems, Waltham, MA). The mRNA levels of the target genes were normalized with GAPDH housekeeping gene levels and compared between patients with and without POAF.

To assess atrial structural remodeling, left atrial (LA) antero-posterior diameter and LA area were evaluated by transthoracic echocardiography and compared between the two groups. Histological analysis of a right atrial appendage sample was also performed. The sample was rapidly immersed into 10% formalin and processed for paraffin histology. Deparaffinized tissue sections were then stained with Masson's trichrome and Hematoxylin-Eosin and the presence and the degree of inflammatory infiltration, subepicardial and interstitial adipose tissue, and perivascular and interstitial fibrosis were evaluated semiquantitatively using a 0–3 scale, where 0 indicated the absence, 1 indicated the presence of mild, 2 indicated the presence of moderate, and 3 indicated the presence of severe changes in the analyzed parameters. The results were compared between patients with and without post-CABG AF.

To evaluate cardiac autonomic remodeling, heart rate variability (HRV) analysis was performed using the continuous 24-h 12-lead ECG recordings obtained prior to CABG. The standard deviation of normal RR intervals (SDNN), the root mean square of successive differences between normal heartbeats (RMSSD), and the proportion of successive RR intervals that differed by >50 ms (pNN50) were analyzed as time domain parameters. The low-frequency (LF) and the high-frequency (HF) components of the HRV spectrum and the ratio between the two (i.e., LF/HF) were analyzed as frequency domain parameters. All parameters were compared between the AF and no-AF groups.

All patients were prospectively monitored for clinically-relevant postoperative events up to the time of discharge. The occurrence of in-hospital death, stroke, and kidney and/or liver dysfunction were recorded as major post-CABG complications. Liver dysfunction was defined as a ≥3-fold increase in liver enzymes and kidney dysfunction as an increase in plasma creatinine with ≥0.5 mg/dL compared to baseline values. Surgical wound infection and postoperative hemorrhage were noted as minor complications. The duration of inotropic therapy and the length of hospitalization were also recorded.

Categorical data are expressed as absolute numbers and percentages. Due to the relatively small number of patients included in the study, all continuous variables are presented as median and interquartile range and the Mann-Whitney test was applied for between-group comparisons regardless of distribution. Fisher's exact test was used for comparison of categorical data. Spearman's rank correlation method was used to assess correlations between the electrophysiological parameters and the occurrence of post-CABG AF. Multiple logistic regression analysis was used to assess the ability of POAF to independently predict post-CABG length of hospital stay and occurrence of post-CABG renal dysfunction. The models were adjusted for age. A p-value < 0.05 was considered statistically significant. All parameters were measured in a blinded manner and all data were analyzed using MedCalc version 12.4.3.0 (MedCalc Software, Ostend, Belgium).

Multi-approach assessment of ECG, electrophysiological, and molecular data failed to reveal any markers of preexisting atrial electrical remodeling in patients who developed post-CABG AF. P-wave duration, amplitude, and fragmentation were not significantly different between the two groups neither in lead II nor in the Lewis leads (all p >0.05; Table 2). Pre-CABG atrial arrhythmic burden and mRNA expression levels of the six studied genes encoding for atrial ion channels were also similar between the two groups (all p >0.05; Table 2).

Viable myocytes were only found in six of the 30 atrial appendage samples used for the electrophysiological study; two were collected from AF and four from no-AF patients. Hence, direct comparison of these parameters between the two groups could not be performed. However, when correlations were ascertained, none of the evaluated action potential parameters was significantly correlated with the occuerence of POAF (all p >0.05).

None of the analyzed echocardiographic parameters (Table 3) was significantly different in patients with AF compared with those who did not develop post-CABG AF (all p >0.05). However, patients who developed POAF had more important subepicardial adipose tissue compared to those who did not (p = 0.02; Table 3). Although POAF patients were older than their non-arrhythmic counterparts, there was no significant correlation between age and the amount of subepicardial adipose tissue (p = 0.92). None of the other analyzed histological parameters was significantly different between the two groups (all p >0.05; Table 3). Figure 2 depicts typical histological findings in patients with and without post-CABG AF.

Figure 2

None of the frequency domain parameters of the HRV spectrum (Table 4) was significantly different between the two groups (all p >0.05). However, RMSSD, a marker of parasympathetic modulation, and mean heart rate were both significantly higher in the group of patients who developed POAF than in their nonarrhythmic peers (both p = 0.03; Table 4).

A single patient included in the study died during follow-up, who did not present POAF until the time of death. Despite the development of POAF, the duration of inotropic treatment was similar in the AF versus the non-AF groups (0 [0–3] vs. 1 [0–2] days, p = 0.81). Occurrence of postoperative liver dysfunction (18.18% vs. 15.78%, p = 1.00), minor complications (54.54% vs. 31.57%, p = 0.45), and stroke (9.09% vs. 0.00%, p = 0.36) was also similar regardless of the presence or absence of post-CABG AF. However, occurrence of post-CABG renal dysfunction (45.45% vs. 15.78%, p = 0.04) and length of hospital stay (17.00 [12.00–20.00] vs. 12.50 [10.75–14.75] days, p = 0.04) were significantly more important in patients who developed POAF than in those who did not. After adjusting for age, POAF remained an independent predictor of length of hospital stay (p = 0.04), but not of renal dysfunction (p = 0.07).

Our study has a number of limitations. This is a single-center study and, as a result, the number of patients included in the study was rather small. More accurate data could be obtained by increasing the number of patients included in future studies. Also, the study population was relatively homogeneous and, although our results are in line with those of previous studies, extrapolation of these results to the entire population of patients undergoing CABG should be done with caution. The electrophysiological study was particularly affected by the small sample size, since viable atrial myocytes were only found in six of the 30 atrial tissue samples, precluding adequate electrophysiological data comparisons. However, compared to other similar studies, in which the success rate of the electrophysiological study has usually been around 10%, the success rate in the present study was considerably higher (i.e., 20%). Remodeling of the right atrial appendage, evaluated in the present study, may not fully reflect histological and/or functional changes in the left atrium. However, recent evidence has shown the existence of proarrhythmic changes in the right atrial appendage in various clinical conditions associated with AF³⁴. In addition, although the incidence of POAF was relatively high in the present cohort, the population evaluated in our study was relatively healthy (i.e., relatively low SYNTAX and CHA₂DS₂VASc scores) and young (i.e., mean age 65.27 ± 5.85 years). As a result, preexisting (structural, electrical, and autonomic) remodeling was quite limited, as reflected by the data obtained in both AF and no-AF patients. This low remodeling burden is likely to have contributed to the lack of significant differences observed in most of the studied parameters between the two groups. Studying larger populations, with higher cardiovascular risk and more important cardiovascular disease burden could bring important additional information in this regard. Thus, above all, the present study should be viewed as a basis for future larger-scale studies.