The Role of a Standardised Two-session Peri-transfer Acupuncture Protocol in Frozen Embryo Transfer for Women with Recurrent Implantation Failure: A Retrospective Cohort Study.

In vitro fertilisation (IVF) is one of the most widely used procedures in assisted reproductive technology (ART), a broad field encompassing various methods designed to help individuals or couples with infertility achieve a successful pregnancy.[ 1 ] IVF involves the fertilisation of an oocyte outside the body, typically in a laboratory setting.[ 2 ] In a standard IVF cycle, the woman undergoes controlled ovarian stimulation to produce multiple oocytes, which are then retrieved and fertilised with sperm in vitro . The resulting embryos are cultured for several days before transfer into the uterus with the aim of achieving pregnancy. IVF may be performed using a woman’s own oocytes and her partner’s sperm or, when indicated, donor oocytes, donor sperm or donor embryos.[ 3 ] Despite substantial advances in IVF techniques, many patients continue to experience difficulty achieving pregnancy, particularly those with recurrent implantation failure (RIF). RIF remains a major clinical challenge in ART because it is associated with repeated treatment failure, emotional distress and increased financial burden. Historically, RIF was defined as repeated failure to achieve implantation or pregnancy despite the transfer of high-quality embryos into a normal uterine cavity.[ 4 ] However, definitions have varied across studies. More recently, the European Society of Human Reproduction and Embryology (ESHRE) updated its recommendations and defined RIF as the failure to achieve a clinical pregnancy after at least three transfers of good-quality embryos, particularly in younger individuals.[ 5 ] This updated definition reflects the multifactorial nature of RIF and the limited effectiveness of available strategies. The underlying causes of RIF are complex and may involve uterine, embryonic, immunologic and hormonal factors.[ 6 ] Although several interventions, including pre-implantation genetic testing (PGT), endometrial receptivity testing, immune modulation and hormonal support, have been investigated, their clinical utility remains controversial.[ 7 ] Consequently, women with RIF often continue to have suboptimal IVF outcomes, underscoring the need for further research into adjunctive treatment strategies. Acupuncture has increasingly been used as a complementary therapy in reproductive medicine. In the broader literature, several mechanisms have been proposed to explain its potential role in improving IVF outcomes, including modulation of uterine and ovarian blood flow, reduction of stress and anxiety, regulation of autonomic nervous system activity and possible effects on endometrial receptivity.[ 8 9 ] These proposed mechanisms have generated growing interest in acupuncture as an adjunct to embryo transfer, particularly in women with repeated implantation failure. However, acupuncture protocols vary substantially across studies, and not all regimens are designed to directly target these mechanisms. Previous studies have explored the potential role of acupuncture in IVF, but the reported findings have been inconsistent.[ 10 ] This inconsistency may be related to heterogeneity in study populations, IVF stage, treatment timing, treatment intensity, acupoint selection, control interventions and outcome definitions. Importantly, many previous studies have evaluated heterogeneous infertility populations rather than specifically focusing on women with RIF, and relatively limited data are available on simplified peri-transfer acupuncture protocols in frozen embryo transfer (FET) cycles. According to the Population, Intervention, Comparison and Outcome (PICO) framework, this study aimed to determine whether, in women with RIF undergoing FET, a standardised two-session peri-transfer acupuncture protocol, compared with no acupuncture, was associated with improved live birth outcomes. Clinical pregnancy rate and ongoing pregnancy rate were evaluated as secondary outcomes. Our hypothesis was that this simplified protocol might confer benefit through relaxation-related effects proposed in previous studies; however, given the heterogeneity of the existing literature, its efficacy and routine applicability remain uncertain. Therefore, the present study evaluated the effectiveness of this specific protocol rather than assuming a general benefit of acupuncture in women with RIF.

The participant selection process is shown in Figure 1 . A total of 2586 women were screened for eligibility from the medical records reviewed during the study period. After exclusion of duplicate cases, donor cycles, cycles involving PGT, cases with abnormal uterine conditions and other ineligible records, 1969 women fulfilled the study criteria and were included in the final analysis. Only one FET cycle per patient was analysed. Of these, 1110 women chose adjunctive acupuncture and were included in the acupuncture group, whereas 859 women declined acupuncture and were included in the non-acupuncture group. Flowchart for case eligibility. PGT = Pre-implantation genetic testing Baseline demographic, clinical and embryo-related characteristics were generally comparable between the acupuncture and non-acupuncture groups [ Table 1 ]. The diagnosis of RIF was based on the patients’ previous embryo transfer history, whereas the embryo-related variables shown in Table 1 describe the index FET cycle included in the present analysis. The mean age was 33.2 ± 4.9 years in the acupuncture group and 33.5 ± 5.0 years in the non-acupuncture group. Similarly, BMI, AMH level and prolactin level were comparable between the two groups, with no statistically significant differences observed. The distribution of infertility causes was also broadly similar across the groups, with male factor infertility representing the most common category in both the groups. In addition, neither the number of transferred embryos nor the number of transferred good-quality embryos differed significantly between the two groups, indicating that the baseline and embryo-related characteristics were broadly balanced. Baseline characteristics of the included individuals *Data are presented as mean±SD, Independent t -test, **Data are presented as n (%), Chi-square test. BMI=Body mass index, AMH=Anti-Mullerian hormone, SD=Standard deviation The embryo-related variables reported in Table 1 refer to the index FET cycle included in the analysis. Therefore, a value of 0 for the number of transferred good-quality embryos indicates that no embryo transferred in that cycle met the predefined criteria for good quality, although embryo transfer was still performed. Pregnancy outcomes remained comparable between the acupuncture and non-acupuncture groups after adjustment for age, BMI, AMH level, number of embryos transferred and number of transferred good-quality blastocysts [ Table 2 ]. For the primary outcome, the AOR for live birth was 0.91 (95% CI: 0.75–1.09, P = 0.312). The AORs were 0.97 (95% CI: 0.81–1.17, P = 0.783) for clinical pregnancy and 0.95 (95% CI: 0.79–1.15, P = 0.600) for ongoing pregnancy. Overall, these findings indicate that the standardised two-session peri-transfer acupuncture protocol was not significantly associated with improved reproductive outcomes. Pregnancy outcomes between the acupuncture and non-acupuncture groups Data are presented as n (%). Adjusted for age, BMI, AMH level, number of embryos transferred and number of transferred good-quality blastocysts. AOR=Adjusted odds ratio, BMI=Body mass index, AMH=Anti-Mullerian hormone, CI=Confidence interval To address the potential confounding effect of advanced maternal age, a sensitivity analysis was performed after excluding women aged 40 years and above [ Table 3 ]. After this exclusion, 1728 women remained in the analysis, including 986 in the acupuncture group and 742 in the non-acupuncture group. The results remained materially unchanged. In women aged under 40 years, the AOR was 0.99 (95% CI: 0.82–1.21, P = 0.936) for clinical pregnancy, 0.96 (95% CI: 0.79–1.17, P = 0.686) for ongoing pregnancy and 0.91 (95% CI: 0.75–1.10, P = 0.332) for live birth. Sensitivity analysis excluding women aged 40 years and above Sensitivity analysis after excluding women aged 40 years and above. Adjusted for age, BMI, AMH level, number of embryos transferred and number of transferred good-quality blastocysts. AOR=Adjusted odds ratio, BMI=Body mass index, AMH=Anti-Mullerian hormone, CI=Confidence interval

In this retrospective cohort study, we evaluated the association between a standardised two-session peri-transfer acupuncture protocol and pregnancy outcomes in women with RIF undergoing FET. The study was conducted using the updated ESHRE 2023 definition of RIF, namely failure to achieve a clinical pregnancy after at least three transfers of good-quality embryos. A total of 1969 women were included, making this one of the larger studies on this topic. After adjustment for age, BMI, AMH level, number of embryos transferred and number of transferred good-quality blastocysts, clinical pregnancy, ongoing pregnancy and live birth rates remained comparable between the acupuncture and non-acupuncture groups. Because advanced maternal age may adversely affect embryo competence and reproductive outcomes in the absence of pre-implantation genetic testing for aneuploidy (PGT-A), we additionally performed a sensitivity analysis excluding women aged 40 years and above. The results remained materially unchanged, suggesting that inclusion of this subgroup did not substantially influence the overall findings. Taken together, these findings indicate that the two-session acupuncture protocol evaluated in this study was not significantly associated with improved reproductive outcomes. However, our results should be interpreted within the context of the specific intervention tested. Importantly, this study was designed to evaluate a simplified, standardised peri-transfer acupuncture protocol used in routine practice at our centre, rather than to assess the overall efficacy of acupuncture as a broad therapeutic concept for all women with RIF. Several previous studies have reported favourable effects of acupuncture on pregnancy outcomes in women undergoing ART, including those with RIF. For example, Junjian et al . reported improved live birth and ongoing pregnancy rates in women with RIF who received acupuncture.[ 11 ] However, their protocol differed substantially from ours, as treatment was initiated 3 months before FET and administered 2–3 times per week for 20 min per session. In addition, their definition of RIF, which required at least two implantation failures, differed from the ESHRE 2023 recommendation. Similarly, Gan et al . reported improved biochemical and clinical pregnancy rates following acupuncture,[ 12 ] but their intervention was initiated immediately after the previous failed cycle and continued until the day before embryo implantation, with treatments administered 3–4 times per week for 30 min per session. A meta-analysis has also suggested a potential benefit of acupuncture in improving pregnancy outcomes.[ 10 ] Nevertheless, the overall quality of the included studies was limited, and those conclusions should therefore be interpreted with caution. In contrast, our findings did not demonstrate a significant benefit of acupuncture. This is consistent with the randomised controlled trial by Andersen et al ., which found no significant difference in pregnancy outcomes between women who received acupuncture and those who did not.[ 13 ] Similarly, two meta-analyses reported that acupuncture may appear beneficial when compared with no adjuvant treatment, but that this apparent benefit is not maintained when acupuncture is compared with sham controls.[ 14 15 ] Together, these findings suggest that although acupuncture may exert certain physiological or non-specific effects, its role in improving reproductive outcomes remains uncertain. One possible explanation for the inconsistency across studies is the marked heterogeneity in acupuncture protocols. Previous studies have differed substantially in treatment frequency, treatment duration, timing of intervention and the type of acupuncture used, including manual acupuncture, electroacupuncture, auricular acupuncture, non-invasive acupuncture and protocols combined with gonadotropin-releasing hormone agonists.[ 9 10 15 16 ] This variation makes direct comparison difficult and limits firm conclusions regarding the overall efficacy of acupuncture in this setting. In addition, a meta-analysis suggested that acupuncture administered during controlled ovarian hyperstimulation and at higher treatment intensity may be associated with better reproductive outcomes.[ 17 ] One study reported that acupuncture performed 25 min before embryo transfer improved IVF outcomes, whereas an additional session after transfer did not provide further benefit.[ 18 ] In the present study, by contrast, we evaluated a brief peri-transfer protocol intended primarily to promote relaxation and reflect routine clinical practice at our centre. Several previous studies reporting favourable outcomes used multi-session regimens over a longer treatment period. It is therefore plausible that a more prolonged or intensive intervention could have different physiological effects from a short peri-transfer relaxation-oriented approach. Accordingly, our findings should be interpreted as applying specifically to this simplified two-session protocol rather than to all acupuncture strategies used in reproductive medicine. Differences in study populations may also contribute to inconsistent findings. Variations in age distribution, ovarian reserve, infertility characteristics and the definition of RIF may all affect outcomes and reduce comparability across studies. In addition, some studies have included relatively small cohorts or non-standardised populations, which may further contribute to inconsistent results. It is also possible that acupuncture does not directly address the principal causes of implantation failure in this population, where embryo competence, endometrial receptivity and immunologic factors may play more central roles. Although multiple practitioners delivered acupuncture in our study, all followed an identical predefined protocol, which helped minimise variability in treatment delivery within the centre. A major strength of this study is the relatively large sample size, which enhances the precision of the estimates and strengthens the analysis compared with many previously published studies in this field. In addition, the study was conducted using the ESHRE 2023 definition of RIF, which improves the clinical relevance and methodological consistency of the cohort. The use of a standardised peri-transfer acupuncture protocol and inclusion of a non-acupuncture comparison group also provide useful insight into the performance of this specific intervention in routine practice. However, several limitations should be considered. First, the acupuncture points were selected primarily to promote relaxation rather than to directly target infertility-related mechanisms, which may have influenced the observed outcomes. Second, the protocol was limited to a short peri-transfer treatment window, and the relatively brief duration of exposure may have reduced its potential impact. Third, acupuncture is a widely accepted complementary therapy in our setting, and participants were not systematically screened for acupuncture use before starting their IVF cycle. As a result, prior acupuncture exposure could not be excluded and may have acted as a potential confounder. Fourth, group allocation was based on patient preference rather than randomisation, which may have introduced selection bias. In addition, acupuncture was self-funded by the patients, which may have further influences. In routine clinical practice, a commonly reported reason for declining acupuncture was fear of needles; however, these reasons were not systematically categorised in the retrospective dataset. Therefore, residual confounding and self-selection bias cannot be excluded. Another limitation is that no formal a priori sample size calculation was performed. Thus, although all eligible cases within the predefined study period were included, the study may still have been underpowered to detect small differences between the groups. In addition, pregnancy follow-up data in our centre were partly obtained through telephone contact with patients, and some important cycle-specific factors influencing FET outcomes, particularly endometrial thickness, were not available in the retrospective dataset. Consequently, some outcomes were based on patient self-report, which may be subject to reporting inaccuracy, and complete follow-up information was not available for all cases. This may also explain the incomplete availability of miscarriage and multiple pregnancy data. Finally, maternal and neonatal outcomes were not assessed, limiting the evaluation of long-term reproductive outcomes. Future studies should investigate acupuncture protocols involving more sessions, earlier initiation in the treatment course and ideally delivery by practitioners with expertise in infertility care. Randomised controlled trials incorporating appropriate sham controls would be particularly valuable in distinguishing specific treatment effects from placebo or non-specific effects. The findings of this study contribute to the ongoing discussion regarding the role of acupuncture as an adjunctive therapy in women with RIF undergoing IVF and FET. Although no significant difference was observed between the acupuncture and non-acupuncture groups, the study highlights the importance of carefully evaluating complementary interventions before incorporating them into routine practice. An important distinction should be made between protocol reproducibility and routine clinical applicability. The present study did not evaluate a comprehensive infertility-focused acupuncture regimen but rather a simplified relaxation-oriented peri-transfer protocol used in routine clinical practice. At the centre level, the intervention was reproducible because all practitioners followed an identical predefined regimen. However, broader routine application of this simplified protocol remains uncertain given the heterogeneity of the existing literature and the absence of improved reproductive outcomes in our cohort. Because RIF is a complex and multifactorial condition, further research using standardised definitions, clearly described acupuncture protocols and consistent outcome measures is needed. Greater methodological consistency across studies would improve comparability and provide clearer guidance for clinicians seeking to optimise ART outcomes.

In this retrospective cohort, a standardised two-session peri-transfer acupuncture protocol was not associated with improved clinical pregnancy, ongoing pregnancy or live birth rates in women with RIF undergoing FET. These findings do not support the routine use of this simplified acupuncture protocol as an adjunctive treatment in this population. TTTT and HPT conceptualised the study. DTPL, LTL and TNKH collected the data and performed the statistical analysis. HPT investigated the results. TTTT, TTDH and NTT interpreted the data and drafted the manuscript. All authors read and approved the final version of the manuscript. There are no conflicts of interest. The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request. Artificial intelligence tools were used to assist with language editing and grammar checking during the preparation of this manuscript. All scientific content, analysis and conclusions were conducted and verified by the authors.

The study protocol and all materials received approval from the Ethics Committee of Hung Vuong Hospital, Ho Chi Minh City, Vietnam (Code: CS/HV/24/36). All patients provided written informed consent for clinical treatment at our centre. For this retrospective study based on existing medical records, the requirement for additional written informed consent for research participation was waived by the Ethics Committee in accordance with institutional policy. The study was conducted in accordance with the ethical principles of the Declaration of Helsinki. In this retrospective cohort study, medical records from Hung Vuong Hospital, Ho Chi Minh City, Vietnam, between January 2019 and December 2023 were reviewed. A total of 2586 women aged 18–45 years were screened. Women were eligible if they had a history of at least three failed embryo transfers, with at least one good-quality embryo transferred in each previous cycle, in accordance with the ESHRE 2023 recommendations for RIF. No formal a priori sample size calculation was performed; instead, all eligible cases identified during the predefined study period were included in the analysis. To avoid non-independence of observations, only one FET cycle per patient was included. Donor cycles, surrogacy cycles and cycles involving PGT were excluded. In addition, women with abnormal uterine conditions were excluded. Participants were informed about the optional use of acupuncture as a complementary treatment and were free to choose whether to receive it. The cost of acupuncture was covered by the patients themselves. Those who elected to receive acupuncture comprised the acupuncture group, whereas those who declined comprised the non-acupuncture group. All included women underwent blastocyst-stage FET. Demographic and clinical baseline characteristics collected from the medical records included age, body mass index (BMI), anti-Müllerian hormone (AMH) level and prolactin level. In addition, embryo-related characteristics were recorded, including the number of embryos transferred and the number of transferred good-quality blastocysts. Embryo quality was assessed according to the Istanbul consensus criteria using the routine morphological evaluation system applied at our centre. Good-quality embryos were defined in accordance with these consensus-based criteria. The reported embryo-related variables refer to the index FET cycle included in the present analysis. The primary outcome was live birth rate, while clinical pregnancy rate and ongoing pregnancy rate were evaluated as secondary outcomes. Clinical pregnancy was defined as the presence of a viable foetal heartbeat confirmed at 7–8 weeks of gestation. Ongoing pregnancy was defined as a pregnancy continuing beyond 20 weeks of gestation. Live birth was defined as the delivery of a live-born infant. Participants began taking 2 mg of oral estradiol (Progynova, Bayer, Germany) twice daily starting from days 2 to 3 of their menstrual cycle. The dosage was increased by 4 mg every 5 days, with a maximum daily dose of 16 mg. Throughout the following 14–16 days, a transvaginal ultrasound was performed to monitor the thickness of the endometrium. Luteal phase support was started once the endometrial thickness reached between 8 and 14 mm and displayed a triple-line pattern. The acupuncture points used in this study were selected primarily to promote relaxation during the peri-transfer period, in accordance with routine practice at our centre, rather than to directly target specific infertility-related mechanisms. Acupuncture was performed in two sessions for each individual undergoing FET. The first session took place before the embryo transfer and included auricular acupuncture at Shenmen (TF4), Sympathetic (Jiaogan-AH6a), Liver (CO12) and Kidney (CO10), alongside body acupuncture at Baihui (GV20), Guilai (ST29), Guanyuan (CV4), Qihai (CV6) and Xuehai (SP10). Needle retention for this session was 25 min. Following the embryo transfer, the second session of acupuncture was administered, which involved body acupuncture at Taichong (LV3), Zusanli (ST36) and Sanyinjiao (SP6), with a needle retention of 25 min. Pulse rate and blood pressure were monitored before and after both acupuncture sessions to assess physiological response. All acupuncture procedures were performed by licensed traditional medicine practitioners with at least 5 years of clinical experience. Although multiple practitioners were involved, all followed the same predefined protocol to ensure procedural consistency within the centre. This standardised two-session peri-transfer protocol reflects routine clinical practice at our centre and differs from traditional multi-session regimens; therefore, its external generalisability and routine clinical applicability should be interpreted with caution. Data analysis was performed using R software (version 4.0.2; R Foundation for Statistical Computing, Vienna, Austria). Descriptive statistics were used to summarise the baseline characteristics of the two groups. Continuous variables are presented as mean ± standard deviation, whereas categorical variables are expressed as frequencies and percentages. Differences between the acupuncture and non-acupuncture groups were assessed using the independent t -test for continuous variables and the Chi-square test for categorical variables. Multivariable logistic regression analysis was performed to estimate adjusted odds ratios (AORs) and 95% confidence intervals (CIs) for clinical pregnancy, ongoing pregnancy and live birth. The models were adjusted for age, BMI, AMH level, number of embryos transferred and number of transferred good-quality blastocysts. P < 0.05 was considered statistically significant.