This is a parallel designed, exploratory randomized controlled trial with 7:3 allocation to intervention group (contains Group Implantation and Group Medication) versus control group. The study is designed to investigate central nervous mechanism of menopausal panic disorder patients with treatment of acupoint catgut embedding, reveal neural activity and amino acid metabolism of PD patients, and internal relationship with clinic curative effect by the use of fMRI and 1H-MRS. The report follows the National Standards of P.R (GB/T 21709.10-2008) manipulations of acupuncture and moxibustion—Part 10 Acupoint Catgut Embedding. The protocol adheres to the SPIRIT 2013 statement; further details are in the Additional file 1. The trial will be done in Bao’an TCM Hospital of Guangzhou University of Chinese Medicine.
There are 3 ways to recruit participants: 1 Recruit from clinic patients of 3 cooperative hospitals. We will gather each hospital’s director of acupuncture department and gynecology department and call a seminar focusing on how to recruit participants. Then we will send assistants to each department to help screen participants, in the meanwhile put up a post about this study to attract potential candidates. 2 Recruit via broadcasting, TV and newspapers, etc. We will give a brief description about our expecting participants and provide free examinations and treatments to those who meet the standards in these advertisements. 3 Attract suitable participants by introducing some basic knowledge about menopausal physiology, menopausal pathology and its treatment in community service.
All the subjects are menopausal syndrome specialist clinic patients from menopausal syndrome clinical research base of Guangdong Traditional Chinese Medicine and Affiliated Bao’an TCM Hospital of Guangzhou University of Chinese Medicine. And healthy female volunteers who are in normal child-bearing age and from the same region act as the control group. Study subjects will receive oral and written information and must sign an informed consent form before enrolment. The inclusion and exclusion criteria are presented in Table 1.
In a baseline questionnaire, information will be obtained about age, occupation (physical/non-physical), education level, last menstrual period, previous use of acupuncture, expectation towards acupoint catgut embedding. Moreover, PDSS, PASS, HAMA Score, SLC-90 Score must be completed at baseline (week 0). Thereafter, the study subjects will be randomized.
Random numbers are generated by PEMS 3.1, and made into random cards. Then the cards are sealed in impellucidus envelopes, kept by designated personnel. There is a one-to-one correspondence between the registration of clinic patients and the serial number of the envelope which contains the random number. Based on the number in the envelope each patient gets, these 70 patients are grouped into Group Implantation and Group Medicine at random, and each group contains 35 samples. In the meanwhile, healthy female volunteers who are in normal child-bearing age are chosen as the control group.
Since it is unachievable to apply blind method to experimental subjects of two groups above and the experimenters, this study propose to apply blind method to result measurers and data statisticians.
Intervention and setting
The study will take place in Affiliated Bao’an TCM Hospital of Guangzhou University of Chinese Medicine. Patients of Group Implantation get one-time acupoint catgut embedding every 2 weeks, while patients of Group Medication take Alprazolam (Guangxi Southpearl Pharmaceutical Co., Ltd.) with size of 0.4mg tablet orally, 1 tablet a time, 3 times a day. There are 3 months study period and 1 month follow-up period for each group. Meanwhile, there is no intervention towards healthy female volunteers who are in normal child-bearing age in control group, except one index examination the day they join the study. A more detailed process flow can be seen in Fig. 1.
Medical material information
The needles will be sterile disposable No.8 injection needles size 0.8 × 38TW LB (Zhejiang KangKang Medical-Devices Co., Ltd.). The core needles will be sterile disposable No.28 acupuncture needles size 0.35×50mm (Rui Qier) used after their pinpoints rubdown. The catgut will be absorbable surgical suture (medical catgut), size 3/0 (Shanghai Pudong Jinhuan Medical Products Co., Ltd.).
Main acupoint: BL23, SP6, RN4.
If it’s a liver-kidney yin deficiency case, add BL18.
If it’s a kidney yang deficiency case, add ST36 and BL20.
In each treatment, RN4 is a required acupoint, and the rest acupoints can be used in alternation of left/right side.
Location method: according to National Standards of P.R <Location of Acupoints> promulgated by the State Bureau of Technology Supervision. Specific acupoints locations are as followed. (Table 2)
Acupoint Catgut Embedding
Operating method (implantation with needles): according to National Standards of P.R (GB/T 21709.10-2008) manipulations of acupuncture and moxibustion—Part 10.
- Cut the 3/0 catgut into 1.5cm segment and soak them in disinfectant.
- During the treatment, patients take prone position to get back-shu point implantation and supine position to get RN4, SP6 and ST36 implantation. Based on location method above, disinfect routinely at acupoint and its skin around with Anerdian, take one self-made embedding needle, stick the core into needle tubing, and then pull out the core about 2cm, implant one 1.5cm sterile catgut into the front of the tubing, and tighten or pinch the local skin around the acupoint with left thumb and forefinger, stick the needle into the acupoint rapidly with the right hand, and the lift and down the needle until acquire de qi. Push the core and withdraw the tubing slowly, and finish implanting the catgut deep down the acupoint. Hold pressure for a moment after withdraw the needle, check for exposure of catgut out of skin and bleeding, cover the wound with a piece of gauze or band-aid for 1-2 days.
- Direction, angle and depth of implantation: BL23, SP6, RN4 are required vertical puncture into the acupoint for about 0.8-1.0 cun (approximately 20mm-25mm); ST36 is required to vertical puncture into the acupoint for about 1.0-1.5 cun (approximately 25mm-40mm); BL20 is required vertical puncture into the acupoint for about 0.5-0.8 cun (approximately 13mm-20mm); BL18 is required oblique puncture towards the spine into the acupoint for about 0.5-0.8 cun (approximately 13mm-20mm).
fMRI examination procedure
The MRI examination will be performed to show changes of brain’s function directly and timely through the blood oxygen level imaging which can offer visualized proof. The parameters of the MRI (Siemens) protocol are as followed: high-resolution three-dimensional T1 weighting brain volume (Bravo) sequence scanning with repetition time (TR) of 8.16 ms, echo time (TE) of 3.18 ms, flip angle of 7°, field of view (FOV) of 256 × 256 mm², voxel size of 1 × 1 × 1 mm³, number of slices of 180 and slice thickness of 1.0-mm. The subjects will be asked to keep quiet and their eyes closed. Meanwhile, they need to empty their minds but avoid sleep. The researcher will check whether the participants are awake during the scanning procedure.
Outcomes will be evaluating using multiple indexes. The primary outcomes are measured by the following therapeutic indexes: frequency of panic attack during observation period (3 months) and during follow-up period (1 month), Panic Disorder Severity Score (PDSS), Panic Associated Symptoms Score (PASS), and HAMA Score. The secondary outcomes are measured by the following therapeutic indexes: SLC-90 Score, Chinese Menopause Rating Scale Score. And there are two Biomarker Indexes: fMRI and 1H-MRS scans taken pretherapy and post-treatment, free amino acid and E2 of peripheral blood tested pretherapy and post-treatment.
fMRI will focus on the activity of Amygdaloid necleus and the connections between fear center and fear network to reflect the changes of brain’s function of menopausal panic disorder patients before and after treatment. 1H-MRS will test Glutamate-GABA level of these brain tissues to explore the relations between neural activity and amino acid metabolism.
All study subjects will be measured by all those indexes except PASS. Frequency of panic attack, Panic Disorder Severity Score (PDSS), HAMA Score, SLC-90 Score, and Chinese Menopause Rating Scale Score will be measured every 4 weeks, and those two Biomarker Indexes will be tested two times: in week 0 and week 12.
Based on preliminary experiments, it is obtained that after treatment the kuppermann index (MI) of Group Acupoint Catgut Embedding drops 15.90±2.71, while Group Medication drops 13.84±1.87, and the population standard deviation S=2.12, δ=2.06; given α=0.05bilateral, β=0.1, power of test 1-β=0.9. Based on formula, n=2x[(tα+ t2β)x s/δ]x[(tα+ t2β)x s/δ], calculating with trial and error method, and use table look-up: tα, ∞=1.96, t2β, ∞=1.282, then we obtain n=23.66≈24, then according to t table: t0.05(24×2-2)=2.0129, t0.2(24×2-2)=1.3002, then substitute it into the former formula, we obtain n=23.25≈24, the sample size has plateaued, and the attempt stop. Which means each group requires 24 samples, and two groups require 48 samples in total(n=48), with the consideration of maximum lost rate of follow up is 20%, we conclude that the total sample size is 60. According to HAMA (which contains 14items), the sample size is 14×5=70. Take two parts above into consideration, the sample size is determined as 70.
Image data processing: using methods like independent component analysis (ICA) and function binding, applying professional softwares such as MRIcro for windows, Xjview 8.1, SPM 8 and Matlab 5.1.
Statistical analysis technique: applying statistical test to difference of prior-treatment and posttreatment, examining the curative effect difference within groups and among groups of all curative effect index. We adapt two-sided test for all statistical tests, it’s considered that the tested differences are statistically significant if P less than or equal to 0.05, while if P greater than or equal to 0.05, then the tested differences are not statistically significant. Details are as follows:
Measurement data: adapt t-test to run comparison among groups, when it doesn’t comply with the normal distribution, we change into Wilcoxon rank-sum test. We run paired t-test towards difference of prior-treatment and posttreatment, and we change into Wilcoxon rank-sum test while it doesn’t comply with the normal distribution.
Enumeration data: we use Chi-square test, calibration Chi-square test, and Fisher exact method to run comparison among groups.
Ranked data: we use Wilcoxon rank-sum test to run comparison among groups and signed rank sum test to run comparison within groups.
In the research, some adverse reactions may occur. We will monitor every patient with adverse reaction. If patient feel any discomfort during interviews, please contact and consult your research physician in time.
There are a few common responses after the acupoint catgut embedding:
- Local reaction: Mostly some sterile inflammations with inflamed hot pain occur within 5 days. Some cases may be more sever, like a small amount of milky-white exudation at the implantation spot caused by fat liquefaction because of catgut stimulation. Circumstances above should not require any special treatment.
- General reaction: Some patients may experience temperature rising after the treatment 4 to 24 hours, which mostly around 38℃. It will fade away in 2 to 4 days by itself. Usually hemogram of each patient may rise in different levels, and it will come back to normal in 3 to 5 days.
There are a few adverse responses after the acupoint catgut embedding:
- Pain: Apply hot compress if there is pain at implantation acupoint.
- Secondary infection: Lack of strict asepsis and well protection of the wound could cause secondary infection, which mostly occurs some inflammatory symptoms like local swelling, progressive pain, and fever 3 to 4 days after the implantation. Apply hot compress and anti-infective treatment would help.
- Nerve injury: It usually caused by incorrect operation or excessive stimulation or carelessness. It can be avoided with careful operation.
- Hemorrhage: It usually caused by puncture on the vessels or excessive stimulation, in which case pressure dressing on the puncture point would stop the bleeding.
- Catgut allergy: Local itching or swelling or fever can be remitted by antianaphylaxis treatment.
If any of above adverse responses occurs, participants should contact their research physicians in time, and they will offer corresponding treatment and advice. If there are any notable findings that may affect the continuation of this research, subjects or their legal representatives will be notified.
Patients need to inform their family or close friends that they are participating in a clinical research, and they can pay attention to adverse responses presented above. If they have any questions about this study, subjects can offer the contact information of their research physicians.
There are some risks of Alprazolam: Some patients will use alprazolam in the research (medication group). Alprazolam is one of Benzodiazepines, and its principle effects are antianxiety and antidepressant. Common adverse reactions are as followed:
- a) Somnolence, vertigo, and weakness etc, large dose may cause dystaxia, tremor, uroschesis or jaundice.
- b) Alprazolam is addictive. Drug withdrawal may cause withdrawal symptoms after using for a long period, showing as emotional or melancholy.
- c) A few patients may suffer dry mouth, inattention, hyperhidrosis, palpitation, constipation or diarrhea, blurred vision, hypotension.
If participants have any questions about having Alprazolam, they could consult their research physician.
All the investigators have been trained well to follow the trial’s procedure and acupuncturists are instructed to give the acupuncture treatment merely. Any other treatment or counseling is forbidden. To ensure the quality of the study, the clinical monitors will arrange specialized staffs to check the process of the trial and document the details of medicine use. Additionally, monitors nominated by the principal investigator will check the accuracy and validity of the original data which should be uploaded in time and accompanied with text summaries. Regular meetings will be held to handle with the difficulties and problems emerging during the study.