The study protocol was approved by the institutional ethical committee. Written informed consents were obtained from all participants prior to enrollment in this study.
This was a prospective randomized study performed on 50 treatment-naïve adult patients with chronic HCV infection—confirmed by a polymerase chain reaction (PCR) test to have HCV genotype 4—presenting to the specialized virology clinic at our institution from February 2019 to August 2019.
All patients were treated according to the national protocol for the treatment of chronic HCV infection which is in line with the recommendations of the European Association for the Study of the Liver [9]. Patients received sofosbuvir 400 mg daily and daclatasvir 60 mg daily for 12 weeks.
Patients were excluded from the study if they had any of the following criteria: Age less than 18 years old; co-administration of beta blockers, calcium channel blockers, amiodarone, or ivabradine; history of coronary artery disease; history of heart failure; more than mild valvular stenosis or more than moderate valvular regurgitation; atrial fibrillation; history of arrhythmias; uncontrolled hypertension; co-infection with hepatitis B virus, HIV, or bilharziasis; an estimated glomerular filtration rate less than 30 ml/min; previous treatment for HCV infection; current liver cirrhosis; anemia; thyrotoxicosis; or active inflammation.
Patient interviews
Patients were interviewed for a detailed history and physical examination. Presence of any of the exclusion criteria was assessed. General and local abdominal examination aimed to detect manifestations of chronic liver diseases and exclude liver cirrhosis. Local cardiac examination aimed to exclude undiagnosed cardiac conditions.
Venous blood samples were obtained from all patients to perform quantitative PCR of HCV antibody prior to treatment and following the completion of treatment, in addition, to routine baseline investigations which are a prerequisite for patient selection for suitability for DAA treatment according to the national protocol. These include aspartate transaminase, alanine transaminase, total bilirubin, serum albumin, alkaline phosphatase, prothrombin time, international normalized ratio, serum creatinine, glycated hemoglobin, complete blood count, and alpha fetoprotein. Additionally, abdominal ultrasound was performed to detect the presence of liver cirrhosis, splenomegaly, or ascites.
Cardiac investigations
The following was performed before the start of treatment and after completion of the 12-week regimen (for practical purposes, the follow-up cardiac investigations were performed during any of the last 3 days of the treatment regimen):
Surface ECG: to assess heart rate, rhythm, measure PR interval, and estimate the corrected QT interval duration (QTc).
PR interval duration was measured from the beginning of the P wave to the beginning of the QRS complex. QT interval was measured from the beginning of the QRS complex to the end of the downward slope of the T wave crossing of the isoelectric line, and then, QTc was calculated according to the formula QTc = QT/square root of R–R interval [10, 11].
24-h ECG monitoring: This was performed using a NORAV Medical NR-302 device which is a three-channel Holter monitor (NORAV Medical GmbH, Wiesbaden, Germany) with five ECG cables connected in designated locations on each patient’s chest.
Device attachment: to ensure a proper recording with minimal artifacts, all patients were prepared by hair shaving (if present) at the site of electrode attachment; the skin was cleaned and gently abraded to improve conductivity and to hold the electrodes in place for the whole duration of the recording. Cables were then connected to the electrodes at the pre-defined sites, and the required identifier data was input to the device.
Device data analysis: all recordings were analyzed by an experienced electrophysiologist blinded to the patient information. The following variables were assessed: (1) minimum heart rate; (2) maximum heart rate; (3) average heart rate; (4) HRV time-domain and frequency-domain measures; (5) significant pauses (longer than 2.5 s); (6) sinus tachycardia episodes (where sinus heart rate exceeds 100 bpm); (7) sinus bradycardia episodes (where sinus heart rate is less than 60 bpm); (8) number of premature atrial contractions (PACs); (9) number of premature ventricular contraction (PVCs); (10) episodes of atrial fibrillation, supraventricular, or ventricular tachycardias.
HRV time-domain measures:
The following time-domain measures were assessed to estimate the variability in the measurements of the time period between successive heartbeats [12,13,14]:
- SDNN which is the standard deviation of NN intervals measured in milliseconds.
- SDANN which is the standard deviation of the average NN intervals for each 5 min segment of the 24-h HRV recording measured in milliseconds.
- RMSSD which is the root mean square of successive RR interval differences measured in milliseconds.
- HRV triangular index which is the integral of the density of the RR interval histogram divided by its height.
HRV frequency-domain measures:
Frequency-domain measures were assessed to estimate the distribution of power in four frequency bands where power is the signal energy found within a frequency band. The European Society of Cardiology and the North American Society of Pacing and Electrophysiology task force divided heart rate oscillations into ultra-low-frequency (ULF), very-low-frequency (VLF), low-frequency (LF), and high-frequency (HF) bands. The following measures were assessed [12,13,14]:
- ULF power which is the absolute power of the ULF band (≤ 0.003 Hz).
- VLF power which is the absolute power of the VLF band (0.003–0.04 Hz).
- LF power which is the absolute power of the LF band (0.04–0.15 Hz).
- HF power which is the absolute power of the HF band (0.15–0.4 Hz).
where absolute power for the previous measures is calculated in milliseconds squared divided by cycles per second [12,13,14].
Statistics
Data from the 50 patients were collected, coded, and statistically analyzed using IBM Statistical Package for Social Science (IBM Corporation, Armonk, NY, USA). Continuous variables that passed normality test were expressed as mean ± standard deviation and analyzed using two-tailed Student’s t test. Categorical variables were expressed as number and percentage and analyzed using chi-squared test. The level of significance was defined at a p value less than 0.05. All baseline parameters were statistically homogenous and normally distributed giving a study power of more than 80.