Effect of acupuncture on endometrial receptivity in women with thin endometrium: study protocol for a single-centre, randomised, sham-controlled trial.

Thin endometrium is clinically defined as endometrial thickness (EMT) below the threshold thickness for successful embryo implantation. When follicles gradually mature during the natural cycle, EMT typically reaches 8–13 mm with increasing oestrogen secretion, creating an optimal microenvironment for embryo implantation. 1 A positive linear relationship has been observed between the EMT measured on the day of human chorionic gonadotropin (HCG) administration and the clinical pregnancy rate (CPR) in the in vitro fertilisation and embryo transfer (IVF-ET) cycle. 2 3 Thin endometrium occurs in approximately 2.4% of IVF cycles. 4 At present, thin endometrium is often defined as EMT of less than 7 mm when the maximum follicle diameter is greater than 18 mm. 5 According to the 2019 Canadian Fertility and Andrology Society guidelines, 6 EMT <8 mm or 7 mm may negatively impact pregnancy and live birth rates in fresh or frozen IVF-ET cycles, respectively. The pathogenesis of thin endometrium involves complex mechanisms. Established aetiological factors include repeated intrauterine procedures (such as artificial abortion and curettage), uterine infection, clomiphene citrate administration and primary biological factors. 7 8 Current strategies for thin endometrium primarily focus on exogenous oestrogen supplementation to stimulate endometrial proliferation, 9 along with adjunctive therapies like low-dose aspirin 10 or sildenafil citrate 11 to enhance uterine perfusion. However, current evidence supporting the efficacy of these interventions in improving EMT remains inconclusive, with no demonstrable improvement in pregnancy rates observed in patients with thin endometrium undergoing embryo transfer. 12 Emerging interventions such as intrauterine infusion of granulocyte colony-stimulating factor, 13 14 autologous platelet-rich plasma infusion 15 and stem cell therapy 16 show promise but lack robust evidence and carry elevated risks. Thus, new strategies for thin endometrium remain an urgent clinical priority. Acupuncture has emerged as a promising treatment for women with thin endometrium. Clinical evidence indicates that acupuncture could enhance endometrial morphology by increasing EMT 17 and type A endometrium proportion 18 and improve uterine artery blood flow. 19 The therapeutic mechanisms may involve angiogenic activation via VEGFR2-dependent PI3K/AKT-ERK cascades, 20 pinopode maturation with concomitant upregulation of αvβ3/HOXA10/HBEGF receptivity markers, 21 and oestrogen-regulated glandular proliferation via secretory enhancement and steroid receptor coordination. 22 Nevertheless, the current evidence supporting acupuncture’s efficacy in thin endometrium management remains limited. This randomised sham-controlled clinical trial aims to compare the effects of acupuncture versus sham acupuncture on EMT in women with thin endometrium.

This is a single centre, patient blinded, randomised controlled trial (RCT) conducted at the Acupuncture and Moxibustion Hospital of China Academy of Chinese Medical Sciences. A total of 120 women with thin endometrium will be randomly assigned to receive acupuncture or sham acupuncture in a 1:1 ratio via a centralised computer system. The trial flow is shown in figure 1 . Participants will be diagnosed with thin endometrium on meeting either of the following criteria: EMT <7 mm on HCG administration day during controlled ovarian stimulation cycles. 4 EMT <7 mm on luteinizing hormone (LH) surge day or when leading follicle diameter reaches ≥18 mm in natural cycles. Eligible participants must satisfy all of the following: Women aged between 20 years and 40 years with fertility requirements. Meet the diagnostic criteria of thin endometrium. Volunteer to participate in the trial and sign a written informed consent. Participants will be excluded for any of the following: Uterine diseases affecting uterine cavity morphology or endometrial receptivity, including submucosal fibroids, intramural fibroids larger than 3 cm or smaller than 3 cm that affect uterine cavity morphology, untreated endometrial polyps, uterine adhesions, uterine effusion, uterine malformations, ASRM stage III–IV endometriosis, adenomyosis, etc. Severe ovarian dysfunction, such as premature ovarian insufficiency, polycystic ovarian syndrome, etc. Major systemic comorbidities, including severe cardiovascular and cerebrovascular diseases, liver, kidney, malignant tumour, haematopoietic system and psychiatric conditions. Recent endometrial therapies, including acupuncture within 3 months or pharmacological treatment within 1 month. All acupuncture procedures will be administered by certified licensed acupuncturists possessing a minimum of 5 years of specialised clinical experience in gynaecological acupuncture. Acupuncturists will receive training on the standard operating procedure of acupuncture. Disposable sterile needles of different sizes (0.25 mm in diameter and 25 mm or 40 mm in length, 0.30 mm in diameter and 75 mm in length) ( Hwato brand, Suzhou Medical Appliance Factory, Suzhou, China) will be used in the trial. The acupuncture protocol of this trial consists of two groups of acupoints. Group 1 includes GV20 (Baihui), GV24 (Shenting), GB13 (Benshen), CV12 (Zhongwan), ST25 (Tianshu), CV4 (Guanyuan), KI12 (Dahe), EX-CA1 (Zigong), ST36 (Zusanli), SP6 (Sanyinjiao) and LR3 (Taichong). Group 2 includes BL23 (Shenshu), BL33 (Zhongliao) and KI3 (Taixi). These acupoints should be located according to the WHO Standard Acupuncture Point Locations in the western Pacific region. 23 According to the theory of traditional Chinese medicine, these acupoints can work together to tonify the kidney and regulate the Chong and Ren meridians, which help to improve endometrial receptivity. We will use one group of acupoints each time, and alternatively use these two acupoint groups. Acupoints of GV20, GV24 and GB13 will be needled transversely at a depth of 15–20 mm. LR3 will be obliquely inserted in the direction of KI1 (Yongquan) up to 15–20 mm. The remaining acupoints, except BL33, will be perpendicularly needled at a depth of approximately 25–30 mm, while BL33 will be needled into the third posterior sacral foramina with an oblique downward direction at a depth of approximately 50–60 mm. Each acupoint will be required to obtain the sensation of deqi (including soreness, numbness, distention, and heaviness, and other sensations) with small, equal manipulations of lifting, thrusting or twirling of needles. Participants will receive acupuncture for 20 min per session, three sessions per week (ideally every other day) for 12 consecutive weeks. For the sham acupuncture control, superficial needling at non-acupoints will be used. There are also two groups of non-acupoints. Group 1 includes non-acupoints 2.25 cun (half of the distance from mid-hairline to frontal angle) lateral to GV20 and GV24, 1 cun (fold the distance between the two nipples to eight equal parts, and one equal part is 1 cun) lateral to CV12, ST25, CV4 and KI12, 10 mm backwardly lateral to SP6; 10 mm lateral to ST36, and non-acupoint located in the anterior depression at the junction of the third and fourth phalanxes. Group 2 includes the non-acupoints 15 mm lateral to BL23 and BL33, respectively, and the non-acupoint located in the anterior edge of the Achilles tendon horizontally to KI3. All the non-acupoints will be superficially needled for 2–3 mm without requirement of deqi. The sequence, frequency and retention time of acupuncture will be the same as those of the acupuncture group. Throughout the trial, the participants will be treated separately to prevent communication and will be discouraged from receiving other treatments for thin endometrium.

The primary outcome is the EMT at week 12. The thickness of the endometrium is measured on the HCG day in the IVF cycle or when the maximum follicle diameter is ≥18 mm in the natural cycle via transvaginal ultrasound. Secondary outcomes include the EMT at week 24, the proportion of type A and type B endometrium classified by Gonen criteria, 24 Pulsatility Index (PI), Resistance Index (RI) and the ratio of peak systolic to end-diastolic blood flow velocities (S/D ratio) of uterine artery, endometrium blood flow, serum oestradiol concentration, CPR, and the score of the Self-Rating Anxiety Scale (SAS). The endometrial type, PI, RI, S/D ratio of uterine artery and endometrium blood flow will be measured via transvaginal ultrasound at weeks 12 and 24 on HCG day in IVF cycles or when leading follicle diameter reaches ≥18 mm in the natural cycles. Endometrial measurements will be performed by a fixed experienced sonographer using a Mindray DC-80 colour Doppler ultrasound system (Shenzhen Mindray Biomedical Electronics, Shenzhen, China). Transvaginal ultrasound scanning will be conducted with a 6–8 MHz probe. For EMT, the maximum distance between the anterior and posterior endometrial-myometrial junctions perpendicular to the midline echo is measured on the mid-sagittal uterine plane. Endometrial type is classified according to the Gonen criteria (A/B/C types) based on echogenicity patterns. 24 Endometrial blood flow is assessed using colour Doppler flow imaging in low-velocity mode (pulse repetition frequency 0.3–0.9 kHz) to evaluate distribution (peripheral or central) and vascular density grading. When measuring uterine artery blood flow, the uterine artery trunk is identified at the level of the internal cervical os, and spectral waveforms are acquired after adjusting the sampling volume angle to ≤30°. PI, RI and S/D are automatically calculated by the device. Bilateral measurements will be performed. Serum oestradiol levels are measured via peripheral blood sampling on the day of endometrial ultrasound examination. CPR will be followed up until the end of the trial. SAS is a self-rating scale for measuring the presence and severity of anxiety. 25 It consists of 20 items with a 4-point Likert Scale, with a higher score indicating more serious anxiety. In this trial, a validated Chinese version of SAS 26 will be used and its score will be assessed at the end of the third and sixth menstrual cycles. All trial data will be captured via the electronic source record (ESR) system by certified blinded outcome assessors. These personnel will be trained on data acquisition and source documentation prior to study initiation. The ESR system ensures complete data provenance through immutable audit trails with cryptographic verification and maintains full revision histories and transparent audit capabilities. Scheduled retention reminders will be delivered via WeChat or telephone to enhance protocol adherence and optimise follow-up compliance. For subjects discontinuing treatment or deviating from protocols, endpoint data will be actively ascertained whenever feasible, and reasons for discontinuation will be recorded in detail. Adverse events (AEs) associated with acupuncture will be monitored throughout the trial. Common AEs of acupuncture include severe pain, bleeding after needle withdrawal, haematoma, etc. Details of AEs, such as the name of AE, occurrence and terminal dates, severity, relationship with acupuncture and prognosis will be documented in the ESR system. The flow chart and study design schedule are presented in figure 1 and table 1 , respectively. CPR will be followed up until the end of the trial. CPR, clinical pregnancy rate; PI, Pulsatility Index; S/D ratio, ratio of peak systolic to end-diastolic blood flow velocities; RI, Resistance Index; SAS, Self-Rating Anxiety Scale. The primary outcome of this trial is EMT. Current evidence remains limited regarding the effect of acupuncture on EMT in women with thin endometrium. According to a previous study 27 and clinical observation, we assumed the estimated EMT in the acupuncture group will be 8.48 mm after 12 weeks of treatment and the EMT for sham acupuncture will be 7.27 mm. A sample size of 50 participants per group will be needed to provide 80% power with a two-sided significance level of 5%, assuming an SD of 2.13. Allowing for a 20% dropout rate, 120 participants will be recruited with 60 participants in each group. A total of 120 infertile women with thin endometrium will be recruited via posters, WeChat or online advertisements. Prior to enrolment, all participants will be provided with comprehensive explanations regarding the study objectives, procedures, potential benefits and associated risks through standardised informed consent protocols. Eligible participants will be randomly assigned to acupuncture or sham acupuncture in a 1:1 ratio using central randomisation via a computerised system. The scheme of randomisation will be designed and archived by a statistician who will not be involved in the statistical work of this trial. The allocation sequence will be computer-generated. A strict view permission will be set for the central randomisation system. In this trial, acupuncturists will conduct randomisation while remaining unblinded to group allocation. Participants, outcome assessors and statisticians will maintain blinding throughout the study. Although participant blinding will not be formally assessed in this study, sham control methodology employing superficial non-meridian needling has been validated for maintaining blinding integrity in previous acupuncture trials. 28 This study will employ the ESR system for data management. The ESR system will implement rigorous logical verification and range checks for data values to guarantee data accuracy. A single-operator data entry model will be implemented, with strictly enforced personnel access privileges. Statistical analysis will be performed using SAS V.9.4 (SAS Institute). According to the principle of intention to treat, statistical analysis will be conducted on all randomised subjects. Missing data will be imputed using the multiple imputation method. Continuous data will be presented as the mean and SD if they are normally distributed, otherwise they will be presented as the median and IQR. Categorical data will be presented as the number and percentage. Comparisons between groups will be analysed using an independent t test or Wilcoxon rank-sum test for continuous variables, and a χ 2 test or Fisher’s exact test for categorical variables. All statistical tests will be two-sided, and p<0.05 will be considered statistically significant. Given the trial’s limited sample size and the inherently low-risk nature of acupuncture interventions, neither a data monitoring committee nor predefined interim analyses will be implemented in this trial. This study will employ a tiered monitoring framework to ensure protocol adherence, data validity and participant welfare. Primary monitoring will consist of 100% source data verification via the ESR system by investigator-appointed quality assurance personnel. Independent third-party monitors from the institutional clinical trials centre will conduct secondary monitoring via hybrid (on-site/remote) monitoring with 50% random sampling, targeting protocol-specific elements: standardised acupuncture protocols, trial documentation completeness, source data traceability, informed consent validity, etc. None. This clinical trial strictly follows the Helsinki Declaration and relevant Chinese clinical trial research regulations. The protocol has obtained approval from the ethics committee of the Institute of Acupuncture and Moxibustion, China Academy of Chinese Medical Sciences (Approval No S2024-04-23-1). Written informed consent ( online supplemental file 1 ) will be secured from all participants prior to commencement of study procedures. Results will be disseminated through peer-reviewed publications and scientific conferences. Substantive changes will be submitted to the ethics committee and trial registry. Before signing informed consent, the researcher responsible for participant recruitment will provide participants with a complete and comprehensive introduction of this trial, including the study purpose, study procedure, possible benefits and risks, fee, etc. Each patient will be voluntarily joining the trial and can withdraw at any time. In this trial, data collection strictly adheres to the principle of data minimisation, exclusively acquiring information directly pertinent to the research objectives. Comprehensive de-identification procedures are implemented, whereby all direct personal identifiers (eg, names, national ID numbers) are systematically replaced with cryptographically generated unique subject identifiers. The control intervention employed in this trial may possess therapeutic potential; therefore, post-trial compensatory treatment will not be systematically provided to participants. This protocol has been registered at International Traditional Medicine Clinical Trial Registry (ITMCTR2024000053). This trial commenced on 1 July 2024, and is currently in the recruitment phase. The anticipated completion date is 31 December 2025.

Acupuncture has been increasingly used to treat reproductive diseases in recent years. 29 30 Many studies 17 31 indicate the effect of acupuncture on improving endometrial blood flow and thickness of the endometrium with limited evidence. This randomised, sham-controlled trial aims to reveal the effect of acupuncture for thin endometrium with rigorous methodological design. It is among the earliest studies to assess the effect of acupuncture versus sham acupuncture for women with thin endometrium. In this trial, a standardised acupuncture protocol from over 10 years’ clinical practice for promoting infertility will be applied for treatment of thin endometrium. 32 Traditional Chinese Medicine theory emphasises the importance of Kidney Qi and Chong and Ren meridians for female reproduction, which is the main basis for acupoint selection of this study. It should be noted that the deep needling method of BL33 can stimulate the sacral nerves that innervate the uterus and thus promote the growth of the endometrium. Current clinical practice employs multiple acupuncture control modalities, including blank control, 33 34 non-penetrating placebo acupuncture, 35 sham acupuncture with superficial needling on acupoints or non-acupoints, 28 36 and standard treatment control. 37 38 Nevertheless, the establishment of an optimal acupuncture control paradigm remains a subject of ongoing methodological debate. While non-penetrative placebo acupuncture demonstrates theoretical superiority for efficacy evaluation through maximum elimination of physiological effects, its practical implementation faces dual constraints: diminished needling sensation intensity compared with conventional acupuncture due to the absence of dermal penetration, and elevated risk of unblinding in acupuncture-experienced populations. Consequently, superficial non-acupoint needling has been adopted as the control intervention in this study, prioritising blinding feasibility while explicitly acknowledging the potential for residual therapeutic effects inherent in skin-penetrating needling. 39 40 Ultrasound examination of EMT, endometrial pattern, and endometrial blood flow perfusion is a non-invasive procedure commonly used in evaluating endometrial receptivity and monitoring endometrial development. 41 42 There was a positive correlation between EMT and implantation rates. 43 Women with type A or type B endometrium on the day of HCG administration showed significantly higher CPRs than those with type C endometrium. 44 Endometrial blood flow is an important factor in endometrial growth, 45 which directly reflects the microenvironment of embryo implantation. 46 Lower PI, RI and S/D ratio contribute to intrauterine embryo implantation, 47 while higher values of these parameters lead to poor endometrial growth. 48 The serum oestradiol level can reflect the status of the endometrium. 49 In this study, the aforementioned outcomes will be used to comprehensively reflect the effect of acupuncture on thin endometrium. In addition, we will also observe the effect of acupuncture on patients’ anxiety using SAS, since anxiety shows a significant impact on patients’ physical health and CPR. 50 51 This study has some limitations. First, in consideration of the implementation of blinding, we use shallow needling at non-acupuncture points as a sham control which is absolutely not inert but more convenient for the implementation of blinding, and the acupuncturists are not blinded. Second, this study will not focus on the association between EMT and the occurrence of pregnancy or live birth. Third, as a single-centre RCT, the sample size of this study may not be large enough, which will limit the generalisation of our results. In summary, this trial will use a mature acupuncture protocol to treat thin endometrium. We expect that the findings of this study will provide more reliable evidence for the use of acupuncture as a treatment of thin endometrium.