Author s response to reviews Title: Anti-inflammatory and immune regulatory effects of acupuncture after craniotomy: study protocol for a randomized controlled trial Authors: Seung-Yeon Cho (sy.cho@khu.ac.kr) Seung-Bo Yang (sb-ils@hanmail.net) Hee Sup Shin (realeponym@hanmail.net) Seung Hwan Lee (toast2000@hanmail.net) Jun Seok Kho (neurokoh@hanmail.net) Seungwon Kwon (kmdkwon@gmail.com) Woo-Sang Jung (wsjung@khu.ac.kr) Sang-Kwan Moon (skmoon@khu.ac.kr) Jung-Mi Park (pajama@khu.ac.kr) Chang-Nam Ko (kcn202@khu.ac.kr) Seong-Uk Park (seonguk.kr@gmail.com) (happyomd@khu.ac.kr) Version: 1 Date: 02 Nov 2016 Author s response to reviews: We thank the editors and reviewers of Trials for taking their time to review our article. We have made some corrections and clarifications in the manuscript after going over the reviewers comments. The changes are summarized below: Reviewer #1: Suitability of primary outcome - in the background section, the cited studies mostly described association between cytokines and risk of sepsis (e.g. ref 3). The clinical significance of CRP level is not discussed. There is no reported association between CRP and clinical event (e.g. oedema). It appears that the reason for choosing CRP as the primary outcome is due to previous studies finding association between CRP and electroacupuncture. Whether CRP is a good biomarker for swelling or infection should be discussed.
Response: Thank you for your advice. We agree that the clinical significance of CRP levels requires further explanation. CRP rises in response to surgical procedures or infections, and several previous studies have suggested that the CRP level is a predictor of complications after surgery. We have added a discussion in the background section of our manuscript with the relevant references (pages 4-5). Also, the time-point at which the primary outcome is measured should be specified on page 11. Response: Time-points of all the outcome measurements are mentioned in the first paragraph of the assessment section as The serum CRP, erythrocyte sedimentation rate (ESR), TNF-α, IL-1β, and IL-6 will be assessed four times: within 48 h before the surgery, and 2, 4, and 7 days after surgery (page 9) Perioperative antibiotic prophylaxis - it is not stated in the protocol whether routine antibiotic prophylaxis is used. This, of course, will have implications on the risk of post-operative infection. I would suggest adding this information to the protocol. Response: That is a good point. Perioperative antibiotic prophylaxis is generally performed in patients. Therefore, we added this to the protocol (intervention section) as the conventional treatment in the Department of Neurosurgery (page 8). Measurement of acute phase proteins and cytokines - the description of procedures relating to blood sampling is limited. Cytokines often have a short half-life and degrade quickly. There are circadian patterns to levels of circulating cytokines, and also variation in relation to food ingestion. I suggest adding details of the timing of blood sampling together with time-to-analysis information so that this is replicable and readers can assess reliability of results. Response: Fasting blood samples will be drawn at a fixed time in the morning before breakfast. The samples will be immediately stored at -80 C after centrifugation until the analysis. We have added these details in the assessment section (pages 9-10). Thank you for your advice. Sample size calculation - the actual mathematical calculation of sample size is correct. However, it is uncertain how the authors arrived at 3.2mg/dl difference in CRP level as the difference they want to detect. A study in the British Journal of Neurosurgery (Al-Jabi & El-Shawarby 2010. Value of C-reactive protein after neurosurgery: a prospective study. BJN. 24(6): 653-659) reported CRP level peaks on day 2 and that it is different depending on the surgery (tumour debulking=20.1±2.5mg/dl; aneurysm clipping=16.5±0.7mg/dl). With the study design being patient-preference trial, there may be an unequal balance of surgery types between the intervention and control groups. This can bias the results of the trial either way. Furthermore, interpretation of CRP level is seldom based on an isolated time-point. The trend of CRP level provides more information. Therefore, authors should clarify the rationale behind choose 3.2mg/dl and consider analysing trend of CRP level (additional statistical analyses required). Response:
The CRP difference with a standard deviation was calculated based on previous laboratory results of patients with craniotomy in our hospital from November 2015 to March 2016. As you recommended, we think the CRP level profile is also a good outcome. We decided to add it to the secondary outcome measures of our protocol. We focus on the delta difference of CRP levels between two time points as well as the CRP level profile, and not just the absolute peak value. This is the main difference between our study and the study by Al-Jabi and El-Shawarby. In addition, we will also perform sub-analyses according to surgery types. The outcome measures and statistical analyses sections were revised. Thank you (pages 10-11). Timing of intervention - the protocol states that the intervention starts within 48 hours after the operation. In previous studies reporting a reduction in CRP level following acupuncture, the intervention started soon after surgery or intra-operatively (ref 2 and 16). The delay in the current protocol may not produce the hypothesised degree of effect. Also, this window of intervention is large relative to the overall study period of 7 days. Authors should discuss these aspects or state the rationale behind them. Response: This is a pilot study to explore the anti-inflammatory and immune regulatory functions of acupuncture treatment after craniotomy. In addition, it is practically difficult to administer acupuncture treatments during operations. Therefore, we planned the intervention to be started as soon as possible after the surgery, and used the inability to receive intervention within 48 hours of the surgery as an exclusion criterion for our study. Operative and ward care - the usage of operative drains and/or urinary catheter may influence the risk of infection. Patients requiring intensive care post-operatively, again, would have a different risk of infection. Authors should state whether these information are collected and how they plan to analyse the data. Response: Any problems including infections such as urinary tract infection will be recorded as secondary outcome measures. We will analyze these data to investigate whether there is a significant difference between the two groups. Please check the outcome measures and statistical analyses sections (pages 11, 13). Thank you. Unblinded nature of study - it is unclear whether the analyses will be done by someone who is blinded to treatment allocation. I would recommend this to be included into the protocol. Response: Thank you for your comment. Of course, the statistical analyses will be performed by a researcher who is blind to the allocation. We have added this to the protocol in the statistical analyses section (page 13). Exclusion criteria - presumably patients with autoimmune disorder would be excluded. I understand that the medication criterion probably would catch these patients, but it would be clearer if this was stated. Thinking about the risks associated with acupuncture, perhaps the authors should consider whether to exclude patients with pacemaker or ICD, and pregnant patients.
Response: Thank you for your advice. We have updated the exclusion criteria section based on your recommendation (pages 7-8). Hospital-stay & cost-effectiveness - it is not clear whether all patients stay in hospital for at least 7 days. Patients who had surgery may be able to leave hospital before 7 days. The comment on page 14 (last sentence in Discussion) about cost-effective benefits may not stand if patients are staying longer in hospital and the costs implicated by provision of acupuncture (equipment & trained staff). Antibiotic prophylaxis, for example, may be more cost-effective. Response: The duration of hospitalization will not be extended due to this study. If more than one treatment session is missed, the participant will be excluded from the study according to the dropout criteria. Reason for additional antibiotics - it would be sensible to record the reason for use of additional antibiotics. As clinicians reviewing patients on a day-to-day basis cannot be blinded (intradermal needles in situ), they may be less likely to initiate antibiotics treatment in the intervention group, for example. Authors should explain whether there is guidance on when to start antibiotics locally, or is it based on individual clinician's judgement. Response: Additional antibiotic treatment will be conducted by a surgeon when medically necessary according to the SOP that includes scalp infection, epidural infection, pneumonia, urinary tract infection, etc. Allocation will not influence the antibiotic treatment. Gantt chart - minor point about this. The Gantt chart describes up to day 7, yet the participant flowchart describes the eighth day. I would suggest a consistent use of language i.e. postoperative day 7. Response: Thank you for your pointing out the mistake. The 8th day will be revised to post-op day 7 in Fig. 1. Reviewer #2: 1. You said patients who agree to receive additional acupuncture treatment will be allocated to the study group and patients who do not agree to receive acupuncture will be allocated to the control group, I think your design can not prevent risk of bias, such as selection, performance, and attrition. Please provide arguments to prevent such bias Response: This is a small pilot study to explore the anti-inflammatory and immune regulatory effects of acupuncture. Despite our best efforts, there is an unavoidable risk of bias. After the pilot study yields positive results, we will conduct a follow-up study with a more rigorous design. Thank you.
2. There is some discrepancy between the title and the intervention, your manuscript title is effect of acupuncture on inflammation... but intervention is acupuncture, electroacupuncture and intradermal acupuncture. Response: Strictly speaking, the three interventions may be different. However, as electroacupuncture and intradermal acupuncture are kinds of acupuncture treatment, they can both be included in the acupuncture category. Therefore, we simplified the title using the word acupuncture instead of mentioning the three methods. 3. You said study group received only acupuncture, electoracupuncture and intradermal acupuncture treatment, Don't they receive other treatment (such as pain control, prevention and treatment of infections)? I think all patients will receive treatment (such as pain control, prevention and treatment of infections) after craniotomy in neurosurgery. To avoid unbalance of baseline data, the authors should design a detailed basic treatment protocol. Response: The participants in the study group receive acupuncture treatments as well as conventional treatment (stabilizing vital signs, pain control, perioperative antibiotic prophylaxis, or treatment of infections). We mentioned this in the manuscript (page 8). The current medication will be recorded in the CRF (case report form) every day, and we will compare the difference between the two groups related to medication. Thank you. 4. There is some discrepancy between the title and study design. Your title is the effects of acupuncture on inflammation... but i think anti - inflammatory response... is more appropriate Response: Thank you for your advice. We have modified the title based on your recommendation. 5. You use acupoint LI4 LI11 PC6 ST36 GB39 LR3 GV20, please provide the reason to use that acupoint. Response: We selected the acupoints based on previous studies. We have added the relevant references (page 8). 6. This study is a single center, small sample and non-rcts, so this study will lead to a low-level evidence-based medical research. Response: We agree with your opinion. This is a pilot study to explore the effect of acupuncture treatments after craniotomy. A follow-up study with a more rigorous design such as an RCT will be conducted after the successful completion of this study. Thank you. 7. Background should be succint and Method should be explained in detail
Response: Since this is a study protocol paper, and the concept and study design including the inclusion criteria may seem unfamiliar, we have explained the background in detail. We have described the methods in more detail. Please check the revised manuscript. Thank you. 8. In sample size estimation, please provide the data (such as your pilot study or other article) to figure out the sample size Response: The CRP difference with a standard deviation was calculated based on previous laboratory results of patients with craniotomy in our hospital from November 2015 to March 2016. The serum CRP difference was 3.2 mg/dl with a standard deviation of 3.190. We consulted a statistical expert regarding this. Thank you. 9. In statistical analysis, i think you must be advised by a statistician " if the data are not normally distributed, non parametric methods will be used" sentence should be added and you said following the principle of intention to treat i think FAS or PP is more appropriate Response: Thank you for your advice. We agree with your opinion and have corrected ITT to PP (page 13). We hope that this revised manuscript will meet the publication requirements of Trials. We thank you again for your constructive comments. Sincerely, Seong-Uk Park