Effect of acupuncture and its influence on cerebral activity in patients with persistent asthma: study protocol for a randomized controlled clinical trial

The sample size calculation of the neuroimaging study is different from that of classic randomized controlled clinical trials. Power analyses for neuroimaging studies rely on assumptions about blood oxygenation level–dependent (BOLD) signal amplitude, smoothness, brain location, and other factors that render principled a priori designations difficult. It has been suggested that for fMRI studies, a minimum sample size (n = 20) should be used in order to obtain 80% power with an error threshold of 0.002 at a single-voxel level [21]. Considering a 20% dropout rate, the sample size in this study was increased to 24 per group, for a total of 48 participants needed.

After the baseline assessment, the eligible participants will be randomly allocated with an equal ratio to their respective groups. To avoid bias in researchers subjective factors, randomization will be implemented by a clinical information management system (Beijing Bioknow Information Science & Technology Co. Ltd., Beijing, China). When participant-recruiting staff members decide to recruit an eligible patient with asthma, they will send the patient’s name, sex, age, and telephone number to this system online. Then, the randomized result will be delivered to the acupuncturists.

Because of the different acupoints in the two groups, the acupuncturists will not be blinded to group allocation. Patients in the two groups will be separated into cubicles to refrain from communication. Outcome assessors and statistical analysts will be blind to the procedure and the results of randomization, group allocation, and intervention.

Subjects receive regular medical therapy (budesonide/formoterol [Symbicort; AstraZeneca, Cambridge, UK] or fluticasone/salmeterol [Seretide; GlaxoSmithKline, Brentford, UK]) as usual while participating in this study, except that changing the asthma control medication is not allowed during this study. On the basis of regular therapy, patients with asthma randomly allocated will receive two different acupuncture treatments: puncture at acupoints on the lung meridian or acupoints on the heart meridian.

MRI data will be acquired with a 3.0-T MRI scanner (Siemens AG, Erlangen, Germany) at Huaxi Magnetic Resonance Research Center, West China Hospital of Sichuan University, Chengdu, China. All patients will be asked to stay awake, keep their eyes open, and remain still during the fMRI can. The participant’s head should be placed in the head mask, and a sponge will be inserted to strengthen the fixation of the head. The scanning procedure contains a localizer, high-resolution three-dimensional T1-weighted imaging (3D-T1WI), BOLD-fMRI, and a DTI sequence. The 3D-T1WI scanning parameters will be as follows: repetition time (TR)/echo time (TE) = 1900/2.26 ms; slice thickness = 1 mm; slice number = 30; matrix size = 128 × 128; and field of view (FOV) = 256 × 256 mm. The BOLD-fMRI scanning parameters will be as follows: TR/TE = 2000/30 ms; flip angle = 90 degrees; slice number = 30; matrix size = 128 × 128; FOV = 240 × 240 mm; slice thickness = 5 mm; and total volume = 240. The DTI data will be acquired with the following parameters: FOV = 240 × 240 mm; TR/TE = 6800/93 ms; matrix size = 128 × 128; and slice thickness = 3 mm with no gap. Two diffusion-weighted sequences were acquired using gradient values b = 1000 s/mm² and b = 0 with the diffusion-sensitizing gradients applied in 64 noncollinear directions. All images will be checked by a consultant radiologist at West China Hospital of Sichuan University to exclude unexpected brain lesions in recruits.

The primary outcome measurements are the Asthma Quality of Life Questionnaire (AQLQ) [22] for assessing health-related impairment of QoL in adult patients with asthma [23]. AQLQ contains 32 items, including 4 domains: activity limitation, symptoms, mental health, and environmental stimuli. It will take 5–10 min for the assessor to complete the AQLQ with each participant.

The secondary outcome measurements include the Asthma Control Test (ACT), the peak expiratory flow rate (PEFR), forced expiratory volume in 1 s (FEV₁), Montreal Cognitive Assessment (MoCA), Zung Self-rating Anxiety Scale (SAS), and Zung Self-rating Depression Scale (SDS). The ACT is a questionnaire consisting of five questions with a 5-point scale for each question, and it is validated for assessing asthma control [24]. PEFR and FEV₁ are the main and objective monitors for asthma therapy. Furthermore, this trial also selects SAS, SDS, and MoCA as the secondary clinical outcomes to assess the emotional state and general cognitive function for psychological cognition importance in the pathogenesis of the patient’s asthma.

All outcomes will be assessed at baseline and at the end of treatment. An overview of the outcome measurement at different time points is shown in Table 1.

Safety monitoring will be conducted throughout the trial with reporting of adverse events (AEs) and serious adverse events (SAEs) in each participant’s case report form (CRF). All AEs/SAEs will be reported immediately to the study principal investigator and attending physician. Each AE will be recorded separately.

Clinical data will be managed with printed and electronic CRFs. Only outcome assessors have access to CRFs and will perform data double-entry. The researchers will be required to follow the requirements of the CRF and fill in the relevant information in a timely and accurate manner. The evidence-based medicine center of the CDUTCM will be responsible for monitoring the study and data every 3 months and will make the final decision to terminate the trial.

The primary analysis will be conducted on a per-protocol basis. The data analysis will be completed by statisticians who are independent of the research and blind to the group assignments. For nonimaging data, statistical analyses will be conducted using IBM SPSS 23.0 Statistics software (IBM Corporation, Armonk, NY, USA). For clinical information, data will be presented as means with SDs. Student’s t test and the chi-square test will be used to compare group differences at baseline. A paired t test will be used to compare the clinical outcomes within the groups. Analysis of variance and the Kruskal-Wallis test will be used to compare the clinical feature changes between groups. The significance will be set at the 5% level with two-sided tests. The clinical and neuropsychological data, including AQLQ, ACT, MoCA, SAS, and SDS, will be converted to domain z scores for correlational analysis with imaging data.

The MRI data will be preprocessed and analyzed by statistical parametric mapping with SPM12 (http://​www.​fil.​ion.​ucl.​ac.​uk/​spm/​) and the CONN functional connectivity toolbox 18b (https://​web.​conn-toolbox.​org/​) using MATLAB 2014b software (MathWorks, Inc., Natick, MA, USA). The structural MRI data will be analyzed using the VBM toolbox within SPM12. The steps include checking for artifacts, structural abnormalities, and pathologies; image segmenting; normalizing to standard template; and spatial smoothing. The preprocessing steps of fMRI data include slice timing correction; head motion correction; skull stripping using the Brain Extraction Tool BET (https://fsl.fmrib.ox.ac.uk/fsl/fslwiki/BET); coregistration of the anatomical image to the mean functional image; segmentation of the anatomical gray matter, white matter, and cerebrospinal fluid; normalization to the MNI152 standard template; smoothing; and band-pass filtering. After preprocessing, a series of brain activity information, including amplitude of low-frequency fluctuation, group independent component analyses, seed-based functional connectivity, complex network analyses, and dynamic connectivity analyses, will be calculated to investigate the brain response to different treatment. Pearson correlation analysis will be used to assess the association between the changes of cerebral activity features and the improvement of clinical outcomes in each group. Finally, mediation analysis will be employed to detect whether the effect of acupuncture treatment on the symptoms and QoL is mediated through regulating the cerebral function. In the mediation analysis, the acupuncture method is the independent variate (X), the cerebral function is the mediator (M), and the symptoms and QoL alteration are the dependent variates (Y).

In recent years, brain imaging techniques were applied to investigate the physiopathology of asthma and revealed structural and functional abnormalities in brain circuits involved in the pathogenesis of asthma [13‐17, 25]. These studies provided an approach to investigate the potential mechanism of some interventions for asthma control. Since then, a few longitudinal studies have begun to uncover functional alterations in the brain due to treatment for asthma. For instance, Yu et al. [26] investigated the spontaneous brain activity with resting-state fMRI before and after cognitive behavioral therapy (CBT) in adult patients with asthma. They reported that the beneficial effects of CBT on asthma control were associated with the reversed abnormal spontaneous brain activity in the bilateral occipital lobe and sensorimotor cortex.

Central integration has been considered as an essential mechanism acupuncture’s therapeutic effect since the 1970s [27, 28]. In the past 20 years, with the aid of functional brain imaging techniques such as fMRI, positron emission tomography, electroencephalography, and magnetoencephalography [29], investigators have mapped the cerebral regions/circuits/networks participating in acupuncture effects for treating neuropathies [30‐33], gastrointestinal disorders [34‐36], and motor diseases [37, 38]. These studies provided abundant visual evidence for understanding the central mechanism of acupuncture.

Although acupuncture-related neuroimaging studies of patients with asthma have not yet been conducted, the proven central pathological features of asthma and the rich experience of acupuncture neuroimaging studies provide a basis for investigating the central mechanism of acupuncture in the treatment of asthma in this study.

This study is concerned with the relative specificity of the “meridian–viscera correlation” and is expected to provide evidence for the optimization of acupuncture treatment for asthma. In traditional meridian theories, “meridian–viscera correlation” is an essential principle for acupuncture in the treatment of viscera-related diseases [39]. It means that each meridian pertains to some specific viscera and connects with several viscera. When the viscera are affected, the acupoints on the related meridian can be selected for the affected viscera. For example, the lung meridian pertains to the lung, so the acupoints on the lung meridian can be used for lung diseases. The “meridian–viscera correlation” theory has also been proven by modern clinic trials. Our previous RCT demonstrated that, compared with the acupoint on the gallbladder meridian, acupuncture at the stomach meridian showed a better effect in improving QoL and relieving symptoms for patients with functional dyspepsia (FD) [40], indicating that the stomach meridian is the best choice for treating stomach disease. Our later neuroimaging studies [41] also demonstrated that puncturing at the acupoints on the stomach meridian elicited wider and more remarkable positive modulation in the abnormal cerebral function in patients with FD than that seen for acupoints on the gallbladder meridian. So, this study focuses on the difference in efficacy and cerebral responses resulting from different acupuncture interventions and tries to provide evidence for the “meridian–viscera correlation” theory.

The quality control program is the precondition for the reliability of our results. Our previous study indicated that strict quality control plays an essential role in the guarantee of high repeatability in acupuncture neuroimaging studies [42]. Accordingly, we designed a quality control program focused on the homogeneity of participants, standardization of fMRI scans, and strictness of acupuncture procedures to improve the reproducibility and reliability of the results.

First, one of the important considerations in the neuroimaging study is that the neural activity in humans is highly variable. The connectivity topology of the brain can be influenced by potentially confounding factors such as age [43], sex [44], handedness [45], and even mental state [46]. Hence, rigorous inclusion/exclusion criteria at the stage of participant enrollment will be established in this study. For example, patients with asthma will be restricted to ages between 40 and 65 years and to being right-handed to guarantee the homogeneity of the clinical population. Concurrently, the SAS and SDS scores will be recorded to evaluate the patients’ psychological state and exclude patients with severe anxiety and depression.

The sensitivity to subject motion is another challenge in fMRI studies [47]. Even the slightest source of artifact, such as physiological fluctuations (respiration and cardiac fluctuations) as well as head motion, can highly influence the final estimate of connectivity. Hence, the patients will be asked to stay awake, keep their eyes open, and remain still at the stage of the fMRI scan. To eliminate possible head motion, the participant’s head should be placed in the head mask, and a sponge will be inserted to strengthen the fixation of the head.

The limitations of this study lie primarily in the seasonality of asthma attacks [48] and the diversity of routine medical treatment for patients with asthma [49]. Thus, inconsistency of regular treatments for asthma may confound our results.

In summary, this fMRI trial is designed to investigate the clinical and cerebral activity changes evoked by different meridians (the acupoints on the lung meridian vs. the acupoints on the heart meridian) in patients with mild to moderate persistent asthma and to explore the associations between the cerebral activity changes and clinical variable changes. The results of this trial will help to provide the optimal adjunctive therapeutic approach for improving the QoL of adults with persistent asthma and provide visual evidence for the clinical application of acupuncture for asthma management.