The protocol was developed in accordance with Standard Protocol Items for Clinical Trials with Traditional Chinese Medicine 2018[34] and Standards for Reporting Interventions in Clinical Trials of Acupuncture.[35]
Table 1 Study protocol schedule of enrollment, intervention, and assessment
Legend: MRI Magnetic resonance imaging, PSQI Pittsburgh sleep quality index, GSRS Gastrointestinal symptom rating scale, SAS Self-rating anxiety scale, SDS Self-rating depression scale
Study design
This randomized, single-blind, sham-controlled neuroimaging study will be conducted at the affiliated hospital of Chengdu University of Traditional Chinese Medicine (CDUTCM), China. A total of 60 CID patients with GI disorder will be recruited using advertisements on posters and hospital networks. All patients meeting the inclusion criteria will randomly receive either 20 sessions of real acupuncture treatment or 20 sessions of sham acupuncture treatment. A multimodal MRI examination and evaluation will be performed at both baseline and the end of treatment. Additionally, 30 healthy controls (HCs) matched for age, sex, and education level will receive one multimodal MRI scan and a clinical assessment. A flowchart of the trial design is shown in Figure 1, and the assessment schedule is shown in Table 1.
Inclusion criteria
Patients diagnosed with CID are eligible to participate in the study. Inclusion criteria are (1) aged between 18 and 65 years, right-handed, any gender; (2) meets the CID diagnostic criteria in the International Classification of Sleep Disorders-Third Edition (ICSD-3),[36] and meets the criteria for spleen and stomach disharmony syndrome in the Clinical Terminology of Traditional Chinese Medical Diagnosis and Treatment—Syndromes (GB/T 16751.2-1997);[37] (2) Pittsburgh Sleep Quality Index score above 7 (PSQI score >7); (3) not taking any medication or health care products to improve sleep quality and experienced GI diseases at least 2 weeks before study enrollment; (5) has not participated in other clinical trials within the last month; (6) agrees to voluntarily participate in the study and signs an informed consent form.
The HCs must meet the following inclusion criteria: (1) passes the neuropsychological tests and reports good sleep quality; (2) no GI symptoms; (3) all physiological indexes within the normal range following a physical examination, and no previous functional or organic disease or head injury; (4) agrees to voluntarily participate in the study and signs an informed consent form.
Exclusion criteria
Patients with any one of the following criteria will be excluded: (1) any severe conditions of the cardiovascular, cerebrovascular, liver, kidney, and hematopoietic systems; (2) secondary insomnia caused by drugs, cervical spondylosis, or other diseases; (3) history of psychiatric and neurological disorders or head trauma with loss of consciousness; (4) abuse of psychotropic drugs for anxiety and depression or sedative and hypnotic drugs; (5) pregnant, preparing for pregnancy, or lactating; (6) MRI contraindications such as claustrophobia, cardiac pacemaker, defibrillator, heart stenting, metal dentures, or intrauterine device.
The exclusion criteria for the HC are the same as for the patients.
Withdrawal criteria
Participants with any of the following conditions will be withdrawn: (1) complications that affect safety; (2) serious adverse events; (2) unwilling to follow the study protocol for examination and treatment.
Sample size
This is a trial to investigate the mechanism of acupuncture and is based on neuroimaging technologies. The requirements of imaging studies[38] and previous similar studies[39-40] suggest that 12–26 individuals in each group is a reasonable sample size for brain image data analysis. To allow for potential problems such as subject dropout and invalid imaging data owing to head movements, a sample of 30 patients in each group is considered reasonable.
Randomization
The randomization will be conducted using random number lists created by SAS 9.2 (SAS Institute Inc., Cary, NC, USA). Sixty CID patients will be randomly divided into two groups of 30 cases each. A third researcher, who will not participate in the subsequent research process, will place serial numbers, group numbers, and random numbers in opaque envelopes. Each sealed envelope will be renumbered. If participants meet the inclusion criteria, envelopes with the same serial number will be opened in the order of enrollment and the randomized results will be forwarded to the acupuncturists.
Blinding
The blinding of acupuncturists is quite difficult to achieve.[41] Therefore, only patients, outcome assessors, and statisticians will be fully blind in this study.
Interventions
Treatments will be performed by certified acupuncturists with at least 3 years of acupuncture experience. Disposable sterile filiform needles (XingLin acupuncture needle, Φ0.25 × 25 mm/Φ0.25 × 40 mm, Tianjin Yi Peng Medical Instrument Co., Ltd., China) will be used. Both the acupuncture and sham acupuncture treatments will comprise 20 sessions over 4 weeks (five sessions per week).
Real acupuncture group
Patients in the real acupuncture group will receive acupuncture treatment on the acupoints Baihui (DU20), Zhongwan (RN12) and Zusanli (ST36) with disposable sterile filiform needles. All acupoints will be located according to the World Health Organization Standard Acupuncture Locations and are shown in Table 2 and Figure 2. Needles in DU20 will be inserted 0.5–1 cun (a unit of measurement in acupuncture, 1 cun = 25 mm) using horizontal needling. Needles in RN12 and ST36 will be inserted 0.5–1.5 cun using perpendicular needling after skin disinfection. After the needle enters into the skin, needles will be manipulated by twirling, lifting, and thrusting to generate the sensation of de qi in the local tissue. The experience of de qi is characterized by sensations of numbness, heaviness, or distension felt by the patient. All needles will be retained for 30 minutes.
Sham acupuncture group
The sham acupuncture group will receive a superficial skin penetration treatment at sham acupoints (SA). These SA are located near real acupoints (2 cm lateral from DU20, RN12, and ST36) but do not belong to any known meridian and are not conventional acupoints. The SA locations are shown in Table 3 and Figure 2.
Superficial skin penetration at SA, without needle manipulation for de qi, is a common sham acupuncture method used in many acupuncture randomized controlled trials.[42] Patients in this group will undergo an acupuncture procedure similar to the one received by patients in the real acupuncture group.
Table 2 Acupoint locations for the real acupuncture group
Acupoints
|
Locations
|
Baihui (DU20)
|
The intersection of the crown midline and the apex of the ears
|
Zhongwan (RN12)
|
On the upper abdomen, anterior median line, four cun* above the navel
|
Zusanli (ST36)
|
On the outside of the calf, three cun below Dubi (ST35)**
|
Legend: *According to the theory of traditional Chinese medicine, the navel is 8 cun below the xiphoid process; **ST35 is located on the lateral depression of the patellar ligament
Table 3 Sham acupoints locations for the sham acupuncture group
Sham acupoints (SA)
|
Locations
|
SA 1
SA 2
|
2cm to the right of DU20
2cm to the right of RN12
|
SA 3
|
2cm outside of ST36
|
Legend: cm Centimeter
Concomitant medication
Patients will be advised to avoid medication as much as possible during the study. Following medical ethical principles, if patients’ insomnia symptoms do not significantly improve and they cannot tolerate their insomnia, they may temporarily take sleeping drugs. Researchers will record the name, dosage, and time of the drugs taken in detail in the Case Report Form (CRF).
Multimodal MRI data acquisition
MRI examinations will be performed at the MRI Center at the University of Electronic Science and Technology of China using a 3.0T MRI scanner (GE Discovery MR750, USA) equipped with a standard 8-channel head coil. MRI scans will be assessed at baseline and after 4 weeks. Subjects will be told to eat light food; avoid drinking strong tea, coffee, and wine; and avoid strenuous exercise the day before the scan. To prevent head movements affecting the image, foam pads and towels will be used to hold the head. During the MRI examination, all subjects will wear earplugs and be instructed to relax with their eyes closed and keep their heads clear. The scanning procedure produces a three-dimensional T1 image (3D-T1), a blood oxygenation level-dependent fMRI (BOLD-fMRI), a DTI sequence, and a proton 1H-MRS. The 3D-T1 scanning parameters will be as follows: repetition time (TR)/echo time (TE) = 5.988/1.972 ms, slice thickness = 1 mm, slice number = 154, field of view (FOV) = 256 × 256 mm. The BOLD-fMRI scanning parameters will be as follows: TR/TE = 2000/30 ms, flip angle = 90°, slice number = 35, matrix size = 3.75 × 3.75, FOV = 64 × 64 mm, slice thickness = 4 mm. The DTI data will be obtained using the following parameters: FOV = 240 × 240 mm, TR/TE = 6800/93 ms, matrix size = 128 × 128, and slice thickness = 3 mm with no gap. The proton 1H-MRS asymmetric PRESS sequence includes TE1 = 25 ms, TE2 = 85 ms, TR=2 s, and a 2×2×2 cm area of interest on the anterior cingulate gyrus will be selected.[26]
Outcome measures
The outcome assessments will be performed at baseline and after 20 treatment sessions. All outcome assessors will be trained in conducting interviews and performing measurements before the study begins and will follow a standard protocol. An overview of the outcome measurement at different time points is shown in Table 1.
Primary outcome
The primary outcome is the PSQI scale score. The PSQI comprises 19 self-evaluation items, among which 18 items generate 7 components. Each component is graded from 0 to 3, and the cumulative score of each component is the total score of PSQI, which ranges from 0 to 21. The PSQI is the most widely used sleep quality rating scale.[43]
Secondary outcomes
The Gastrointestinal Symptom Rating Scale (GSRS)[44] will be used to measure GI symptoms.
The Self-Rating Anxiety Scale (SAS) and the Self-Rating Depression Scale (SDS)[45] will be used to assess subjects’ emotional state.
Data management
Clinical data will be managed using printed and electronic CRF accessible only by the research team. The CRF is the original record and cannot be changed at will. It will be kept by a researcher of the research team. At the end of the trial, two researchers will enter data from the case report form into a computer. The Evidence-based Medicine Center of the CDUTCM will regularly monitor the trial data.
Data analysis
Clinical data analysis
Statistical analyses of the clinical data will be performed using SPSS 20.0 statistical software (IBM Corporation, Armonk, NY, USA) and supervised by a skilled statistician blinded to group allocation. The Kolmogorov–Smirnov test will be used to test the normal distribution of continuous variables. Continuous normally distributed data will be reported as means and standard deviations; continuous non-normally distributed data will be expressed as medians with interquartile ranges. Categorical data will be presented as frequencies or percentages.
For the demographic and clinical information at baseline, the chi-squared test or Fisher’s exact test will be used for comparison of dichotomous data. The two-sample t-test will be used for normally distributed continuous data or the Mann-Whitney U-test for non-normally distributed data. To compare baseline and post-treatment changes in the same group, we will use the paired t-test. Covariance analysis using baseline data as covariates will be used to compare the effects of different interventions on continuous data. A p-value <0.05 will be considered statistically significant.
Imaging data analysis
The fMRI data will be preprocessed and analyzed using SPM12 (http://www.fil.ion.ucl.ac.uk/spm/) and CONN toolbox 18b (https://web.conn-toolbox.org/) with MATLAB 2014b (MathWorks, Inc., Natick, MA, USA). The sMRI data will be analyzed using the VBM toolbox within SPM12. DTI data will be processed using FSL Software on Linux. MRS data will be preprocessed using the commercially available software LCModel spectral-fitting package (version 6.3-1N; Stephen Provencher, Inc., Oakville, ON, Canada). After standard preprocessing of each imaging modality, the following brain activity information will be calculated to examine the neural response to different treatments: the amplitude of low-frequency fluctuation, group independent component analyses, seed-based functional connectivity, cortical thickness, tract-based spatial statistics, and glutamate. Finally, Pearson correlation analysis will be used to assess the association between changes in multimodal neuroimaging features and improvements in clinical outcomes in each group.
Patient safety
During treatment, any adverse events (AEs) will be monitored and recorded in the CRFs throughout the trial. Important AEs in this study would be needle-related AEs, such as bleeding, hematoma, dizziness, infection, neurological symptoms, and fainting. Patients with mild and moderate AEs will receive symptomatic treatment. Severe AEs will be reported to the research ethics committee within 48 hours.
Patients and public involvement
Patients and the public were not involved in the design and implementation of the study. They were also not asked to advise on the reporting and dissemination of the results.