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  • Enhancing childbirth experience: The synergistic effects of free posit

    Enhancing childbirth experience: The synergistic effects of free posit

    Introduction

    Natural childbirth is widely recognized as a relatively safe delivery method for mothers, offering faster postpartum recovery and effectively avoiding the short- and long-term complications associated with cesarean sections.1–3 However, the labor process for natural delivery is often prolonged, particularly for first-time mothers, and it is difficult to avoid pain caused by uterine contractions during labor. This pain not only impacts the delivery process but may also pose certain risks to maternal and neonatal safety.4–6 Labor pain, one of the primary physiological challenges faced by mothers during childbirth, primarily stems from uterine contractions, cervical dilation, and pressure on the birth canal. Such pain can trigger significant psychological stress in mothers, potentially reducing the efficiency of contractions, prolonging labor duration, and consequently increasing the likelihood of cesarean delivery and other complications.7–9 Although traditional supine delivery facilitates medical monitoring, it has been associated with potential disadvantages, including increased reports of maternal discomfort, compromised hemodynamics (eg, supine hypotensive syndrome), and potential narrowing of the pelvic outlet compared to upright positions, potentially exacerbating maternal discomfort and hindering fetal descent.10–12

    In recent years, non-pharmacological interventions have gained increasing attention in obstetric research and clinical practice as part of a broader movement towards humanized childbirth care.13 Studies have shown that evidence-based, woman-centered labor care not only facilitates smoother delivery but also effectively reduces the risk of adverse outcomes.14,15 Among these interventions, free positioning and mindful relaxation techniques have garnered significant attention for their role in supporting the delivery process. Free positioning during labor transcends the limitations of traditional supine delivery, allowing mothers to choose positions such as standing, squatting, kneeling, or lying on their sides according to their needs and comfort. This autonomy in movement is thought to optimize the pelvic angle (potentially increasing the anteroposterior diameter), facilitate the descent and rotation of the fetus by utilizing gravity, and may reduce soft tissue resistance, thereby potentially reducing delivery difficulties and associated pain.16–18 However, widespread implementation of free positioning faces challenges, including staff training requirements, resource constraints (eg, availability of birthing aids like balls or stools), and adherence to conventional protocols in some settings.14,19 Meanwhile, mindful relaxation techniques (MRTs), often rooted in Mindfulness-Based Stress Reduction (MBSR) principles adapted for childbirth, involve deliberate attention regulation, non-judgmental awareness of present-moment sensations (including pain), breathing regulation, progressive muscle relaxation, and positive emotional guidance.20,21 The core aim of MRTs is to help mothers modulate their psychological and physiological stress responses, thereby reducing the affective and cognitive components of pain perception, alleviating anxiety, and enhancing their overall sense of control and childbirth experience.22,23 Despite documented benefits, the integration of structured MRTs into routine antenatal education and intrapartum care remains variable and is not yet standard practice globally.24,25

    Although previous studies have separately explored the roles of free positioning and mindful relaxation techniques in childbirth, rigorous research evaluating their combined application in natural delivery remains notably scarce. Systematic reviews highlight the independent benefits but lack robust evidence on synergistic effects.26,27 Considering the complementary physiological and psychological effects of these two approaches, a synergistic interaction is theoretically plausible: free positioning primarily addresses biomechanical factors and physiological pain pathways (eg, optimizing fetal position and reducing soft tissue strain), while MRTs target the psychological and neurocognitive aspects of pain processing (eg, reducing fear-tension-pain cycles and enhancing pain coping mechanisms).28,29 This combined approach may lead to greater overall pain reduction and labor efficiency than either intervention alone. Therefore, this study aims to examine the combined intervention’s efficacy in alleviating labor pain and shortening labor duration, thereby providing a basis for further optimization of obstetric care strategies. This investigation is particularly relevant within our institutional context in China, where there is a growing national emphasis on promoting natural childbirth and enhancing maternal satisfaction within the healthcare system,30 yet barriers to implementing comprehensive non-pharmacological approaches persist.

    Subjects and Methods

    Study Subjects

    Study Design and Ethical Considerations

    This retrospective cohort study included pregnant women who delivered at Wuxi Second People’s Hospital between August 2023 and October 2024. The study was approved by the Ethics Committee of Wuxi Second People’s Hospital (Approval No.: 2022–081) and conducted in accordance with the 1964 Helsinki Declaration and its later amendments. Due to the retrospective nature of the study, the requirement for informed consent was waived by the IRB. Data collection employed an “opt-out” method, consistent with national regulations and ethical standards for minimal-risk retrospective research using anonymized data. Potential participants were informed about the study via hospital noticeboards and electronic platforms; those declining participation within two weeks were excluded. No objections were registered.

    All collected data were strictly confidential and anonymized before analysis to protect participants’ privacy. Access to the data was limited to authorized research personnel only, and all data handling complied with institutional and national data protection regulations to ensure data security throughout the study.

    Participants

    After screening electronic medical records against predefined criteria, a total of 120 eligible participants were identified and included in the analysis. Participants were categorized into two groups based on the documented care received during delivery: the research group (n = 60), who received the combined intervention of free positioning and mindful relaxation techniques, and the Control Group (n = 60), who received routine delivery care.

    Inclusion and Exclusion Criteria

    Inclusion Criteria

    Participants were required to meet the following criteria:

    Singleton pregnancy at full-term gestation;

    No absolute indications for cesarean delivery and the ability to undergo natural childbirth;

    Availability of complete and traceable clinical records;

    Normal communication abilities and clear consciousness.

    Exclusion Criteria

    Participants were excluded if they met any of the following conditions:

    Presence of pregnancy complications such as hypertensive disorders of pregnancy or gestational diabetes;

    High-risk pregnancies, including advanced maternal age or multiple pregnancies;

    Abnormal pelvic structure or birth canal deformities;

    Coagulation disorders or cognitive impairments;

    Concurrent cardiac, pulmonary, or other major organ dysfunction.

    Interventions

    Control Group: Routine Delivery Care

    Participants received standard obstetric care per hospital protocol. This included continuous fetal heart rate monitoring and assessment of cervical dilation. Upon reaching active labor (cervix dilated ≥3 cm), participants were transferred to the delivery room. Midwives provided continuous presence and support. Participants were typically guided to adopt supine or semi-recumbent positions as labor progressed and were instructed on breathing and pushing techniques during the second stage. Psychological support included verbal encouragement and physical reassurance (eg, hand-holding, gentle stroking).

    Research Group: Combined Intervention (Free Positioning + Mindful Relaxation)

    Midwives delivering this intervention had completed a standardized 40-hour training program covering: 1) Theory and evidence for free positioning and mindful relaxation; 2) Practical demonstration and supervised practice of all positions and mindfulness techniques; 3) Communication skills for guiding women; and 4) Documentation requirements. Training was based on WHO recommendations22 and FIGO guidelines.31

    Free Positioning: During the active phase of labor (cervical dilation ≥3 cm), midwives introduced and demonstrated a variety of labor positions to the participants. Women were encouraged to move freely and select positions based on their comfort and individual preference, with midwives providing support and assistance for any necessary position changes. Commonly adopted positions included: recumbent positions (semi-recumbent, right lateral, and left lateral), standing positions (standing upright beside the bed while holding onto support), sitting positions (seated on the delivery bed or a low stool with hands supporting and body slightly leaning back), squatting positions (feet apart, supported by holding onto a chair or bed edge, often with midwife assistance), and kneeling positions (kneeling on a soft mat with knees apart and leaning forward on a pillow or blanket). There were no fixed time requirements for any specific posture; changes in position occurred as frequently as desired by the woman or in response to discomfort, allowing for a flexible, real-world application of the intervention.

    Mindful Relaxation Techniques (MRTs): Upon admission to the delivery room (with cervical dilation ≥3 cm), guided mindful relaxation sessions were initiated and offered continuously throughout labor, particularly during contractions. Each session typically lasted 10–20 minutes and was flexibly adjusted to align with the pattern of uterine contractions and the participant’s level of concentration. While maintaining a self-selected labor position, participants were guided by midwives through a structured mindfulness protocol. This included sensory focus, wherein women were provided with a real object (such as an apple or orange) and instructed to observe its color, shape, and texture, touch its surface, and appreciate its aroma. This was followed by internalization, where they were guided to close their eyes, mentally recall the object’s characteristics, and focus attention while clearing the mind of distractions. A calm ambiance was created using continuous instrumental music played at a moderate volume (40–60 dB). During guided imagery, participants were encouraged to use their imagination to mentally explore the object in greater detail, promoting immersive concentration. In the breath regulation phase, midwives guided a transition from shallow, rapid breathing to deep, steady diaphragmatic breathing, synchronized with silent counting from one to ten and back. Finally, in the reflection stage, participants were instructed to open their eyes, then close them again to recall and verbalize their experience. This protocol, adapted from Mindfulness-Based Childbirth and Parenting (MBCP) principles,32 aimed to alleviate maternal stress, enhance emotional and physical relaxation, and improve women’s coping with labor pain.

    Observation Indicators

    In this retrospective cohort study, all observation indicators were extracted from standardized medical and nursing records. To reduce bias arising from inconsistent data collection, especially for subjective measures (eg, pain or psychological states), only data recorded by trained staff using standardized tools during the clinical process were included. Records with missing or retrospectively added assessments were excluded to enhance data reliability. The specific indicators evaluated were as follows:

    Duration of Labor

    Labor duration was segmented into the first, second, and third stages, with total labor time subsequently calculated. These data were obtained from partogram records maintained by midwives in real time during delivery. Comparative analysis was conducted between the intervention and control groups to evaluate differences in labor progression.

    Pain Levels

    Pain intensity during labor was evaluated using a multidimensional framework based on the Verbal Rating Scale (VRS) recommended by the World Health Organization in 1980. This included three components: (1) the VRS, which classifies pain into four levels—0 (no pain), 1 (mild pain that does not interfere with sleep or daily life), 2 (moderate pain requiring non-narcotic interventions), and 3 (severe pain requiring narcotic analgesia and often accompanied by autonomic symptoms); (2) the Pain Rating Index (PRI), consisting of 15 descriptors, each rated from 0 to 3 (no, mild, moderate, or severe pain), with higher scores indicating greater overall pain perception; and (3) the Visual Analogue Scale (VAS), a 10-cm horizontal ruler labeled from 0 (no pain) to 10 (worst imaginable pain), where participants marked the intensity of their pain. Only assessments recorded by midwives during labor were included in the analysis.

    Perineal Tears

    The degree of perineal trauma was classified based on clinical records according to standard obstetric grading criteria. These included: intact perineum (no laceration), Grade I (superficial tears of the vaginal or perineal mucosa), Grade II (tears involving the perineal muscle layer and/or posterior vaginal wall), Grade III (tears extending to the external anal sphincter), and Grade IV (full-thickness tears involving the rectal mucosa). Grading was performed by attending midwives or obstetricians immediately postpartum and documented in delivery notes.

    Sense of Labor Control

    Maternal sense of control during childbirth was measured using the Labor Agentry Scale (LAS), which comprises 29 items rated on a 7-point Likert scale, yielding a total score between 29 and 203. Higher scores represent greater perceived control and agency during labor. In this retrospective analysis, only LAS questionnaires completed within two hours postpartum and documented in full by trained staff were considered valid.

    Negative Emotions

    Psychological states, specifically anxiety and depression, were assessed using the Self-Rating Anxiety Scale (SAS) and the Self-Rating Depression Scale (SDS), respectively. Each scale includes 20 items rated on a 4-point scale, with a total score of 80. Higher scores indicate more severe emotional distress. In order to ensure consistency, only data collected at standardized time points (within one week before intervention and within 30 minutes postpartum) and documented under supervision were included in the analysis.

    Pregnancy Outcomes

    Pregnancy outcomes were assessed by recording postpartum hemorrhage volume within two hours of delivery and evaluating neonatal health using the Apgar scoring system. The Apgar score includes five parameters—skin color, heart rate, respiratory effort, muscle tone, and reflex response—each rated from 0 to 2, with a maximum total of 10. Scores were categorized as 10 (optimal condition), 7–9 (mild concerns), or <7 (requires immediate intervention). These scores were extracted from standardized neonatal assessment forms filled by neonatal nurses or pediatricians at 1 and 5 minutes after birth.

    Data Analysis

    A priori sample size calculation was performed using G*Power 3.1.25 Based on pilot data (unpublished) and previous studies,6,18 a medium effect size (Cohen’s d = 0.65) for the primary outcome (pain VAS score) was assumed. To achieve 80% power (α=0.05, two-tailed t-test), 60 participants per group were required. This justified the final sample size of n=120 (60 per group).

    GraphPad Prism 8 was used for graphical presentation. SPSS 26.0 was used for statistical analysis. Quantitative data were assessed for normality using the Shapiro–Wilk test and visual inspection of Q-Q plots. Normally distributed data are presented as Mean ± Standard Deviation (SD) and compared using Independent Samples t-tests. Non-normally distributed data are presented as Median (Interquartile Range, IQR) and compared using Mann–Whitney U-tests. Qualitative data are presented as Number (Percentage, %) and compared using Chi-square (χ²) or Fisher’s exact test, as appropriate. Effect sizes are reported (Cohen’s d for t-tests, Cramer’s V for χ²). A two-tailed P value < 0.05 was considered statistically significant. No adjustments for multiple comparisons were made for secondary outcomes, consistent with exploratory analysis in this retrospective design; findings should be interpreted accordingly.

    Results

    Baseline Data

    The control group included 60 participants, aged 25–35 years (Mean ± SD: 28.44 ± 2.63 years); gestational age 37–42 weeks (39.02 ± 0.94 weeks); body weight 50–78 kg (63.15 ± 6.22 kg).

    The research group included 60 participants, aged 25–35 years (28.96 ± 2.58 years); gestational age 37–42 weeks (39.07 ± 1.02 weeks); body weight 50–78 kg (63.74 ± 6.18 kg).

    No significant differences existed in baseline characteristics (P > 0.05, Table 1), confirming comparability.

    Table 1 Comparison of Baseline Data Between the Two Groups (Mean ± SD)

    Labor Duration

    As shown in Figure 1, the research group exhibited significantly shorter labor durations across all three stages compared to the control group. Specifically, the mean duration of the first stage of labor was markedly reduced in the research group (240.69 ± 25.96 minutes) compared to the control group (362.47 ± 31.94 minutes), with a large effect size (Cohen’s d = 4.22, 95% CI: 112.15–131.41, p < 0.001). The second stage of labor was also significantly shorter in the research group (42.32 ± 10.69 minutes) versus the control group (52.14 ± 12.23 minutes), with a moderate to large effect size (Cohen’s d = 0.87, 95% CI: 6.29–13.35, p < 0.001). For the third stage, the research group showed a mean duration of 5.11 ± 2.56 minutes, significantly less than the control group’s 8.74 ± 2.85 minutes (Cohen’s d = 1.35, 95% CI: 2.78–4.48, p < 0.001). Overall, the total labor duration was significantly reduced by approximately 135 minutes in the research group (291.23 ± 28.65 minutes) compared to the control group (426.56 ± 40.69 minutes), with a large effect size (Cohen’s d = 3.86, 95% CI: 121.30–149.36, p < 0.001).

    Figure 1 Comparison of Labor Duration Between the Two Groups.

    Note: *Indicates P < 0.05 compared to the control group.

    Pain Levels

    As shown in Figure 2, pain levels following the intervention were significantly lower in the research group across all assessment dimensions. On the Visual Analog Scale (VAS), the research group reported a mean score of 4.32 ± 1.03, significantly lower than the control group’s 5.23 ± 1.24 (Cohen’s d = 0.80, 95% CI: 0.65–1.17, p < 0.001), indicating a moderate to large effect size. Similarly, the Pain Rating Index (PRI) scores were reduced in the research group (2.49 ± 0.45) compared to the control group (2.94 ± 0.78), with a moderate effect size (Cohen’s d = 0.71, 95% CI: 0.28–0.62, p < 0.001). In terms of Present Pain Intensity (PPI), the research group scored 25.45 ± 2.14, significantly lower than the control group’s 28.77 ± 2.56 (Cohen’s d = 1.42, 95% CI: 2.67–3.97, p < 0.001), reflecting a large effect size. These findings indicate that the combined intervention of free positioning and mindful relaxation techniques was effective in significantly alleviating both the sensory and cognitive-affective components of labor pain.

    Figure 2 Comparison of VRS Scores Between the Two Groups.

    Note: *Indicates a significant difference between the two groups, P<0.05.

    Perineal Laceration

    As illustrated in Figure 3, the distribution of perineal outcomes differed significantly between the two groups (P < 0.05). In the research group, 35.00% (21/60) of participants had an intact perineum, 41.67% (25/60) experienced Grade I lacerations, 21.67% (13/60) had Grade II lacerations, and 1.67% (1/60) sustained Grade III lacerations. In comparison, the control group showed 23.33% (14/60) with an intact perineum, 30.00% (18/60) with Grade I lacerations, 35.00% (21/60) with Grade II lacerations, and 11.67% (7/60) with Grade III lacerations. These findings indicate a higher proportion of intact perineum and first-degree lacerations but a lower incidence of more severe (Grade II and III) tears in the research group, suggesting a protective effect of the intervention on perineal outcomes.

    Figure 3 Comparison of Perineal Laceration Degrees Between the Two Groups.

    Note: *Indicates a significant difference between the two groups, P<0.05.

    Labor Control

    The analysis of Labor Agentry Scale (LAS) scores revealed a statistically significant difference between the two groups. Participants in the research group reported a higher sense of control during labor, with a mean LAS score of 152.41 ± 8.11, compared to 144.22 ± 9.11 in the control group (d = 0.95; 95% CI: 5.42–10.96; P < 0.001). This suggests that the combined intervention notably enhanced participants’ perceived autonomy and empowerment throughout the birthing process. See Table 2.

    Table 2 Comparison of LAs Scores Between the Two Groups (Mean ± SD)

    Negative Emotions

    The analysis of post-intervention psychological outcomes indicated that participants in the research group experienced significantly lower levels of anxiety and depression compared to those in the control group. Specifically, the Self-Rating Anxiety Scale (SAS) scores were 46.23 ± 2.35 in the research group versus 55.98 ± 2.47 in the control group (t = 22.152, P < 0.001; d = 4.10; 95% CI: 8.99–10.23). Similarly, Self-Rating Depression Scale (SDS) scores were 45.22 ± 1.73 in the research group and 54.56 ± 2.14 in the control group (t = 26.291, P < 0.001; d = 4.71; 95% CI: 8.63–10.06). These findings suggest that the combined intervention was highly effective in reducing negative emotional responses during labor. See Table 3.

    Table 3 Comparison of Anxiety and Depression Scores Between the Two Groups ()

    Pregnancy Outcomes

    Postpartum outcomes revealed significant differences between the two groups. The mean volume of bleeding within 2 hours postpartum was markedly lower in the research group (155.89 ± 22.21 mL) compared to the control group (204.58 ± 30.88 mL), indicating improved hemostatic outcomes (d = 1.79; 95% CI: 40.65–57.59; P < 0.001). Apgar scores were slightly higher in the research group (8.44 ± 1.56) compared to the control group (8.05 ± 1.11); however, the difference was not statistically significant (d = 0.28; 95% CI: 0.13–0.73; P = 0.145), indicating comparable neonatal conditions between groups. See Figure 4.

    Figure 4 Comparison of Postpartum 2-Hour Blood Loss and Neonatal Apgar Scores Between the Two Groups.

    Note: *Indicates a significant difference between the two groups, P<0.05.

    Discussion

    This study aimed to evaluate the effects of free positioning combined with mindfulness relaxation techniques on labor pain relief and labor duration reduction. The results showed that this combined intervention significantly shortened the duration of each stage of labor, alleviated labor pain, improved maternal sense of control and emotional status during labor, and reduced postpartum blood loss, demonstrating favorable clinical outcomes. These findings are consistent with multiple previous studies, further validating the value of this integrated intervention model in promoting natural childbirth.

    Firstly, the significant shortening of labor duration is one of the core findings of this study. The intervention group’s average duration of the first stage of labor was markedly shorter than that of the control group (240.69 minutes vs 362.47 minutes), with an overall labor time reduction of approximately 135 minutes and a very large effect size (Cohen’s d = 3.86). This result aligns with the conclusions of Mansfield et al’s systematic review on free positioning facilitating labor progress, which indicated that freedom of movement and position changes could reduce the risk of prolonged cervical dilation and extended second stage of labor.33 The mechanism may be related to multiple factors: free positioning allows the mother to utilize gravity to assist fetal descent, relieve pressure on the birth canal during uterine contractions, facilitate better alignment of the fetal head with the birth canal curve, and reduce birth canal resistance.34 Moreover, alternating between different positions can improve pelvic morphology and blood circulation, helping relax the pelvic floor muscles and thus accelerating labor.35

    In addition, the effect of mindfulness relaxation in alleviating labor pain and improving emotional state may also partly be attributed to the potential regulation of the neuroendocrine system. Although this study did not directly measure neuroendocrine-related indicators, existing literature shows that mindfulness interventions can reduce sympathetic nervous system activity, enhance parasympathetic tone, and regulate the hypothalamic-pituitary-adrenal (HPA) axis reactivity, thereby decreasing the secretion of stress hormones such as cortisol and norepinephrine.36–38 This neuroendocrine balance adjustment helps lower pain sensitivity, relieve anxiety and tension, increase pain threshold, and enhance maternal sense of control and adaptability during labor. In this study, women in the intervention group performed better in subjective pain scores (VAS, PPI, etc)., anxiety and depression scales (SAS/SDS), and labor control scores (LAS), indirectly supporting the plausibility of this mechanism. This dual physiological–psychological pathway suggests that the combined intervention of free positioning and mindfulness relaxation not only improves clinical outcomes but may also have favorable neuroregulatory and mind-body synergistic effects, warranting further exploration at the physiological mechanism level.

    Simultaneously, the intervention group showed a significantly enhanced sense of labor control (LAS scores significantly higher than control), indicating that this intervention model improved maternal autonomy and self-efficacy. A high sense of control during labor is closely related to reduced labor anxiety, decreased frequency of obstetric interventions, and promotion of vaginal delivery.39 This is because free positioning and mindfulness relaxation allow mothers to actively choose comfortable postures and psychological adjustment methods, increasing their mastery over the delivery process and reducing fear and helplessness.40 This result is also supported by studies by Li and Guo, who reported that psychological support and self-regulation strategies during labor significantly improve maternal delivery satisfaction and psychological health.41,42

    This study also observed a significantly lower incidence of severe perineal lacerations (grade II and above) and a notable reduction in postpartum blood loss in the intervention group. Free positioning (eg, semi-sitting, lateral, squatting) can reduce perineal tension, promote natural soft tissue expansion, and decrease the risk of mechanical injury.43 Mindfulness relaxation may contribute by lowering stress responses, improving vascular tone and tissue perfusion, thereby facilitating local repair and hemostasis.44 These findings are consistent with Hughes’ review on perineal protection strategies, which emphasizes the importance of posture adjustment and emotional interventions in reducing perineal trauma and postpartum hemorrhage.45

    In summary, this study confirms that free positioning combined with mindfulness relaxation techniques significantly promotes labor pain relief, shortens labor duration, reduces perineal trauma, and improves maternal psychological state. The mechanisms involve physiological and mechanical optimization (such as fetal descent and pelvic morphology changes), neuroendocrine regulation (reduced sympathetic excitation and enhanced analgesia), and psychological-behavioral enhancement of labor control. The synergy of these mechanisms improves the overall childbirth experience and facilitates smooth natural delivery.

    Limitations

    Although the results show that free positioning combined with mindfulness relaxation techniques have positive effects on labor pain relief, labor duration reduction, emotional improvement, and childbirth experience enhancement, several limitations should be fully considered when interpreting the findings. First, this study is a retrospective cohort design and is limited by the completeness and accuracy of existing medical records, which may introduce information and recall bias. Since randomization and blinding were not performed, there may be uncontrolled baseline differences and selection bias between the intervention and control groups, affecting the rigor of causal inference. Moreover, potential confounders such as maternal social support, fear of childbirth, experience level of birth attendants, and fetal position were not fully controlled, which could partially interfere with the assessment of intervention effects.

    Second, the sample size is relatively small and drawn from a single center, which may affect the representativeness and generalizability of the results. Future studies should validate these findings through multicenter, large-sample, prospective randomized controlled trials to strengthen external validity and causal inference. Additionally, as this study did not include single free positioning or single mindfulness intervention groups, it cannot clearly evaluate the independent contribution of each component nor conclude whether true synergistic effects exist. Therefore, the term “combined intervention effect” should be used cautiously to indicate superior effects compared with routine care but not to prove interaction between interventions.

    Finally, although literature suggests that mindfulness interventions may exert effects via neuroendocrine regulation—such as reducing sympathetic activity, modulating the HPA axis, and decreasing stress hormone secretion36–38—this study did not directly measure neuroendocrine markers, so these remain hypothetical explanations that need further mechanistic research to validate their biological basis.

    Conclusion

    The results of this study preliminarily suggest that free positioning combined with mindfulness relaxation interventions may help shorten labor duration, relieve labor pain, improve emotional state, and enhance maternal sense of control during childbirth, thereby promoting smooth natural delivery. This non-pharmacological, low-cost intervention model has certain potential for clinical promotion, especially in resource-limited or humanized childbirth-focused settings.

    However, given the retrospective design, lack of randomization, blinding, and long-term follow-up, causal interpretations should be cautious. Additionally, the synergistic effects of the combined intervention cannot be separated to clarify individual components’ independent effects. Future research should employ more rigorous prospective randomized controlled trials or factorial design studies to further elucidate intervention mechanisms and explore applicability and sustainability across different populations and labor stages.

    In summary, the current findings provide valuable preliminary evidence supporting non-pharmacological interventions to promote natural childbirth, warranting further exploration and validation in higher-quality studies to assess feasibility and effectiveness for broader application.

    Disclosure

    The authors report no conflicts of interest in this work.

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    30. Young D. What is normal childbirth and do we need more statements about it? Birth. 2009;36(1):1–3. doi:10.1111/j.1523-536X.2008.00306.x

    31. Ayres-de-Campos D, Spong CY, Chandraharan E. FIGO intrapartum fetal monitoring expert consensus panel. FIGO consensus guidelines on intrapartum fetal monitoring: cardiotocography. Int J Gynaecol Obstet. 2015;131(1):13–24. doi:10.1016/j.ijgo.2015.06.020

    32. Zhang D, Tsang KW, Duncan LG, et al. Effects of the Mindfulness-Based Childbirth and Parenting (MBCP) program among pregnant women: a randomized controlled trial. Mindfulness. 2023;14(1):50–65. doi:10.1007/s12671-022-02046-8

    33. Mansfield B. The social nature of natural childbirth. Soc Sci Med. 2008;66(5):1084–1094. doi:10.1016/j.socscimed.2007.11.025

    34. Azizmohammadi S, Azizmohammadi S. Hypnotherapy in management of delivery pain: a review. Eur J Transl Myol. 2019;29(3):8365. doi:10.4081/ejtm.2019.8365

    35. Evans MI, Britt DW, Worth J, et al. Uterine contraction frequency in the last hour of labor: how many contractions are too many? J Matern Fetal Neonatal Med. 2022;35(25):8698–8705. doi:10.1080/14767058.2021.1998893

    36. Vargas-Uricoechea H, Castellanos-Pinedo A, Urrego-Noguera K, et al. Mindfulness-based interventions and the hypothalamic–pituitary–adrenal axis: a systematic review. Neurol Int. 2024;16(6):1552–1584. doi:10.3390/neurolint16060115

    37. Ring HZ, Kern RJH. Zen meditation and the neuro-immuno-endocrine axis. Health. 2024;16(12):1242–1249. doi:10.4236/health.2024.1612086

    38. Pascoe MC, Thompson DR, Ski CF. Metabolism: meditation and endocrine health and wellbeing. Trends Endocrinol Metab. 2020;31(7):469–477. doi:10.1016/j.tem.2020.01.012

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    40. Ayers S. Fear of childbirth, postnatal post-traumatic stress disorder and midwifery care. Midwifery. 2014;30(2):145–148. doi:10.1016/j.midw.2013.12.001

    41. Li L, Zhang P, Qin Z, et al. The effect of holographic Meridian scraping therapy combined with free position on the labor process, perineum lateral resection rate, and delivery outcomes of primiparae. Am J Transl Res. 2021;13(8):9846–9852.

    42. Guo L, Chen L, Jiao Y, et al. Analysis of the effect of free position delivery on the success rate and safety of vaginal trial delivery in patients with scar uterine vaginal delivery. Panminerva Med. 2022;64(4):574–576. doi:10.23736/S0031-0808.21.04478-5

    43. Dénakpo J, Lokossou A, Tonato-Bagnan JA, et al. [Delivery in free position perhaps a solution to change delivery in traditional position in delivery rooms in Africa: results of a prospective study in Cotonou in Bénin]. J Obstet Gynaecol Can. 2012;34(10):947–953. doi:10.1016/S1701-2163(16)35408-1

    44. Veringa-Skiba IK, Ziemer K, de Bruin EI, et al. Mindful awareness as a mechanism of change for natural childbirth in pregnant women with high fear of childbirth: a randomised controlled trial. BMC Pregnancy Childbirth. 2022;22(1):47. doi:10.1186/s12884-022-04380-0

    45. Hughes A, Williams M, Bardacke N, et al. Mindfulness approaches to childbirth and parenting. Br J Midwifery. 2009;17(10):630–635. doi:10.12968/bjom.2009.17.10.44470

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  • Sleep Fragmentation in TcMAC21 Mouse Model of Downs Syndrome

    Sleep Fragmentation in TcMAC21 Mouse Model of Downs Syndrome

    Jacob Tusk,1,&ast; Marina Antonia Salinas Canas,1,&ast; Tarik F Haydar,1,2 Terry Dean1,3

    1Center for Neuroscience Research, Children’s National Hospital, Washington, DC, 20010, USA; 2Department of Anatomy and Neurobiology, Boston University Chobanian and Avedisian School of Medicine, Boston, MA, 02118, USA; 3Department of Critical Care Medicine, Children’s National Hospital, Washington, DC, 20010, USA

    Correspondence: Terry Dean, Center for Neuroscience Research, Children’s National Hospital, Washington, DC, 20010, USA, Tel +1-202-476-6817, Email [email protected]

    Background / Objective: Down syndrome (DS) is the most common chromosomal disorder worldwide, and approximately ¾ of individuals with DS demonstrate multifactorial sleep disturbances, including sleep apnea. As the effects of chromosome 21 triplication are complex, mouse models may provide valuable insights into the causal mechanisms of disordered sleep in DS. Although the recently developed transchromosomic TcMAC21 mouse model offers the closest genetic similarity to human DS, its sleep-wake architecture is unexplored. We hypothesized that TcMAC21 mice would exhibit sleep disruption similar to human DS, specifically with increased wakefulness and sleep fragmentation compared to the euploid controls.
    Methods: Using a non-invasive piezo-electric sleep recording system, we evaluated the sleep-wake architecture in male TcMAC21 (TS, n=9) and euploid (EU, n=9) male control mice under a 12-hour light/dark cycle. Analyzed metrics included: total sleep percentage, bout frequency, and bout length.
    Results: Compared to EU controls, TS mice exhibited a significant reduction in sleep bout duration (− 29.0%, p =  0.02) during the dark phase, with primary effect during the first 8 hours, culminating in an overall decrease in total sleep percentage (− 24.2%, p =  0.04). The light phase did not demonstrate statistically significant changes in total sleep percentage or sleep architecture.
    Conclusion: TcMAC21 mice demonstrated significant sleep fragmentation during the dark phase, potentially reproducing some aspects of sleep disruption in Down syndrome. Interestingly, these findings differed from descriptions of sleep in other DS animal models. Given the high degree of DS gene replication and non-mosaic nature of the TcMAC21 model, it may provide unique insight into the neurologic and anatomic mechanisms of sleep dysfunction in Down syndrome.

    Introduction

    Down syndrome (DS) is the most common chromosomal abnormality in humans and is caused by the presence of an extra copy of human chromosome 21 (Hsa21). This triplication leads to a multitude of clinical manifestations, including sleep disturbances. Sleep disorders in DS are common and multifactorial, with contributors such as obstructive and central sleep apnea and circadian dysrhythmia. Ultimately, DS patients experience poor sleep efficiency and excessive daytime sleepiness.1–6

    Sleep architecture has been studied in multiple mouse models of DS that bear triplication of mouse chromosome regions that are syntenic to Hsa21. While triplication of mouse chromosome 16 (Mmu16), which mirrors a portion of Hsa21, causes fetal lethality,7 other models each demonstrate a unique sleep phenotype. Dp168,9 mice, which include triplication of the entire Mmu16, exhibit decreased sleep in both the light and dark phases.8 Meanwhile, Ts65Dn mice,10 containing a partial Mmu16 and partial Mmu17, show reduced sleep primarily in the dark phase.11 Ts1Cje mice, with a shorter region of triplication on Mmu16, display no baseline sleep differences but delayed sleep rebound after deprivation.11 Finally, the transchromosomic Tc1 mouse,7 which includes a fragmented mosaic of human chromosome 21 (Hsa21) including about 75% of the protein coding genes, shows sleep fragmentation during the light phase and increased sleep latency during the light-dark transition.12 These diverse sleep phenotypes are thought to be driven by each model’s distinct genetic composition, highlighting the importance of accurately recapitulating the human disorder’s chromosomal abnormality in animal models.

    The newly developed transchromosomic TcMAC2113 mouse model offers a significant advancement for DS research. It replicates 93% of the protein-coding genes on Hsa21q, including key genes associated with DS.13 Furthermore, unlike the mosaic nature of the Tc1 model,7 TcMAC21 is non-mosaic,13 ensuring uniform trisomic genetic material across cells and enhancing the reproducibility of phenotypes. However, the sleep architecture of TcMAC21 mice has not yet been described. Given its close genetic resemblance to human DS, this study aims to characterize sleep patterns in TcMAC21 mice to assess their potential as a platform for exploring the etiology and consequences of sleep disruptions in DS. We hypothesized that TcMAC21 mice would exhibit sleep disruption similar to human DS, specifically with increased wakefulness and sleep fragmentation compared to the euploid controls.

    Materials and Methods

    Mice

    All procedures and experimental design were approved by the Institutional Animal Care and Use Committee at Children’s National Hospital (Protocol 30786) and follows National Institute of Health (NIH) and Animal Research: Reporting of In-Vivo Experiments (ARRIVE) guidelines. Our vivarium is maintained at 72°F ±2°F, and the humidity range is 30–50%. Mice had unrestricted access to standard laboratory diet, water, and nesting squares and were maintained under a 12-hour light/dark cycle. The TcMAC21 line was originally acquired from Jackson Laboratories (Strain 035561). As the TcMAC21 line has been reported to have a high variation of fecundity,14 which is anecdotally consistent with our experience, all TcMAC21 females were reserved for breeding purposes and not available for use in this study; a convenience sample of 10 male TcMAC21 (TS) mice and 10 male euploid (EU) littermates was used. Given the light/dark phase-specific findings noted on previous studies of DS mouse models,7–11 an a priori power analysis based on variability in murine sleep noted in our previous experience15 suggested that n=10 would provide adequate power to detect a 10% decrease in 12-hr total sleep time with 95% certainty, and would exceed the numbers in previous sleep reports in DS mouse models.11 Data were collected over 3 separate litters of mice with as many as 8 mice recorded at a time; each recording run included simultaneous recordings of mice of both genotypes, randomly assigned to sleep chambers within a light-tight, sound-proof cabinet (Actimetrics). The analyzed mice were 6.3 ± 1.2 weeks (TS) and 7.1 ± 0.9 weeks (EU) old at the time recording.

    Sleep Recording and Processing

    The use of a non-invasive sleep recording system limits the potential for surgery-related factors (eg recovery from anesthesia, wound healing, inflammatory response) to influence changes in sleep-wake behavior, which is a consideration in the TcMAC21 mice that bear a global transgenic modification. All mice underwent a 5–7 day acclimation period, during which they were individually housed in the piezo-electric chambers (Signal Solutions) with free access to food and water. Up to 8 chambers are housed within a circadian cabinet that would be used for non-invasive sleep recording. After the acclimation period, those chambers would then continuously collect motor activity for 48 hours without disruption. The activity thresholds for distinguishing sleep and wake states were determined using commercially available SleepStats 2020 (Signal Solutions), which has previously shown 89% sensitivity and 96% specificity for wake versus sleep (NREM + REM);16–18 however, this system’s sensitivities/specificities for distinguishing NREM and REM sleep are limited16 and were not utilized in this study. Data were exported as CSV files and analyzed for sleep-wake epochs, with checks for sensor errors or electrical interference. For each mouse, two consecutive 24-hour light/dark (L:D) cycles were analyzed, generating metrics including sleep bout length histograms and percent sleep. Of note, the TcMAC21 mice did bear distinguishable physical characteristics noted previously,13 including abnormal facies and shortened ears, making blinding investigators to mouse genotype impossible as the investigators interacting with the mice were the same as those collecting and analyzing the sleep data; however, the piezosleep sleep analysis system does not require user input to calculate each of these metrics, limiting the possible introduction of researcher bias in the analysis.

    Data Analysis and Availability

    Statistical analyses of comparisons were conducted using Prism (GraphPad version 10.1). Our primary outcomes of differences in total light and dark phase sleep chosen given the previous histories of finding light- and dark-specific changes in Down Syndrome mice.7–11 After applying Grubbs’ extreme studentized deviate testing (α=0.01) to the 24-hr and light/dark phase total sleep times, two mice were identified as statistical outliers and removed from all analyses: 1 EU that slept 43% less than the group average, 1 TS that slept 58% above the group average; at the time of the recordings no mice were observed to have been ill by appearance or gross motor behaviors. For the remaining mice (9 TS, 9 EU), no further outliers were removed from any analyses. Twenty-four hour, light, and dark total sleep percentages were analyzed via unpaired T-tests. Evaluation of sleep architecture (sleep percentage, sleep bout duration, sleep bout frequency) in 4-hour bins were conducted by two-way repeated measures ANOVA followed by comparisons between genotypes (EU vs TS) per bin; false discovery correction for multiple comparisons was conducted as per Benjamini-Krieger-Yekutieli procedure.19

    Results

    TcMAC21 (TS) mice exhibited a statistically significant reduction in 24-hour total sleep percentage relative to euploid (EU) controls (mean ± SD: 42.8 ± 8.3% and 50.1 ± 4.2%, respectively; p = 0.04; Figure 1A). This difference was accounted for primarily by a significant decrease in dark phase sleep (26.6 ± 6.3% vs 35.1 ± 9.6%, respectively; p = 0.041), with a non-significant reduction during the light phase (59.2 ± 12.5% vs 65.1 ± 5.6%; p = 0.21). Because a previous animal DS model demonstrated a sleep phenotype isolated to the first half of the dark phase,12 we next evaluated sleep metrics in 4-hour intervals (Figure 1A). For total sleep percentage, genotype demonstrated a significant effect (p=0.041) while time of day (p=0.10) and their interaction (p=0.62) did not. Pairwise comparisons identified decreased sleep during the first four hours of the dark phase (ZT 1200–1400; p=0.04), but ultimately none were statistically significant after correction for false discovery rate (Figure 1A). Analysis of the light phase suggested only significant effects of time of day (p<0.001) without an effect of genotype (p=0.21) or an interaction between genotype and time of day (p=0.84).

    Figure 1 Comparison of sleep architecture in TS and EU mice. (A) (left) Sleep expressed as a percentage of total time for 24 hour and 12 hour periods. TS mice (magenta) show reduced sleep compared to EU controls (green) when measured over 24 hours, primarily due to a decrease during the dark phase (indicated by horizontal black bars). (right) Sleep expressed as a percentage of 4-hour intervals throughout the 24-hour day. TS mice showed trends towards reduced sleep during the first third of the dark phase (ZT 1200–1600). (B) Mean sleep bout duration for each 4-hour interval throughout the 24-hour day. TS mice (magenta) show reduced sleep bout duration sleep compared to EU controls (green) during the first 8 hours of the dark phase (ZT 1200–1600, ZT1600-1800). (C) Mean sleep bout frequency for each 4-hour interval throughout the 24-hour day. For all panels, Tukey box plots indicate median and interquartile range (IQR) (via box) and minimum/maximum up to 1.5x the IQR above/below the 25%ile and 75%ile, respectively (via whiskers); outliers represented by individual points. Individual ZT’s are marked the center of a four-hour interval. For all experiments, n = 9 for EU, n = 9 for TS. Any statistically significant P-values are detailed.

    We next considered sleep bout duration and frequency in 4-hour bins to characterize the changes in sleep architecture underlying the observed sleep differences. Genotype (TS vs EU) produced a significant decrease in mean sleep bout duration during the dark phase (240.9 ± 81.5 vs 339.2 ± 80.5 s, respectively; p=0.02; Figure 1B), while the factors time of day (p=0.07) and their interaction (p=0.66) did not. Pairwise comparisons revealed significantly decreased mean sleep bout duration in TS mice during the first 8 hours of the dark period (ZT 1200–1600: 189.0 ± 66.0 vs 301.0 ± 108.7 s, p = 0.02; ZT 1600–2000: 214.0 ± 67.2 vs 340.1 ± 132.6 s, p=0.03), without an effect during the last 4 hours (ZT 2000–2400: 376.4 ± 124.7 vs 319.7 ± 207.2 s, p=0.49). Conversely, no significant effects were seen on mean bout length (Figure 1C) during the light phase (genotype p = 0.49, time of day p = 0.12, interaction p = 0.53). Similarly, no significant differences were found in the analyses of the sleep bout frequencies in either the dark or light phases (dark: time of day p = 0.29, genotype p = 0.76, interaction p = 0.59; light: time of day p = 0.45, genotype p = 0.67, interaction p = 0.33).

    Discussion

    The TcMAC21 (TS) mice demonstrated significant alterations in sleep-wake architecture, most notably driven by a decrease in sleep bout duration during the dark phase, the primary active period for mice. As no compensatory changes in bout frequency were observed during the dark phase, the degree of sleep loss over the 12 hours remained significant. This contrasts with sleep behavior during the primary resting (ie light) phase, which may have trended towards comparatively smaller decreases in overall sleep time and sleep bout duration, but did not reach statistical significance. When compared to other DS mouse models (ie Dp16, Ts65Dn, Ts1Cje, Tc1), the dark-phase-specific decrease in sleep of TcMAC21 mice most closely resembles the sleep behavior of the Ts65Dn model. However, Ts65Dn mice also exhibit an extended period of wakefulness during the first 6 hours of the dark phase, unlike the TcMAC21 mice slept for ~21% during ZT 1200–1600; they both demonstrated shortened sleep bouts when they did sleep during the dark phase. Tc1 mice do demonstrate sleep fragmentation similar to TcMAC21 mice, but the effects are primarily observed during the light phase. These differences highlight the variability in sleep phenotypes across DS mouse models, suggesting that sleep disturbances in trisomy 21 are likely polygenic in origin, with different combinations of triplicated genes contributing to varying phenotypic outcomes (summarized in Table 1).

    Table 1 Comparison of Baseline Sleep Architecture Findings Between Mouse Models of Down Syndrome

    The reduction in sleep bout length observed in TcMAC21 mice is consistent with sleep fragmentation. However, the exact cause of sleep fragmentation in these mice remains unknown. As human DS is associated with obstructive and central sleep apneas, it is possible that the TcMAC21 mice could be experiencing a similar phenomenon. We and previous reports of the TcMAC21 mice have noted changes in craniofacial development, including shorter and wider snouts,13 therefore a contribution of altered airway anatomy to disordered breathing during sleep is possible. We predict that future studies employing whole body plethysmography may determine the roles of obstructive or central hypoventilation to the dark phase disturbances we observed. Furthermore, simultaneous incorporation of polysomnography would also improve upon our system’s limitation being unable to differentiate NREM and REM sleep. There are at least two benefits of polysomnography in this mouse model. The first is that it will be necessary to determine if the shortened sleep bouts seen in the TcMAC21 mice prevent normal quantities of REM sleep, similar to human patients.20 Should the murine sleep behavior prove consistent with the human disorder, then the TcMAC21 mouse may be a useful platform for further investigation of the mechanisms of disordered sleep as well as testing new therapeutic strategies. The second is to provide increased sensitivity for changes in sleep that may have evaded detection by our piezoelectric system. For instance, we noted consistent albeit non-significant trends towards less total sleep percentages in the light phase at a smaller magnitude than our dark-phase findings. Use of polysomnography will provide a more definitive characterization of sleep-wake balance during that phase, determining if the sleep phenotypes are truly isolated to the dark phase.

    The TcMAC21 mouse presents an opportunity to dissect the contributions of sleep disruption, itself, to the greater neurodevelopmental pathophysiology in DS. For instance, sleep disturbance in human DS is associated with impairments in expressive language development.21 The TcMAC21 model may be able to address the role of the sleep phenotype, itself, on communicative ultrasonic vocalizations.22 Similarly, the TcMAC21 mice demonstrate overexpression of amyloid precursor protein in the hippocampus as well as significant learning and memory deficits on behavioral testing,13 which may model the increased susceptibility for dementia in human DS.23 However, it is also known that sleep fragmentation, itself, may contribute to this type of pathophysiology.24,25 Using TcMAC21 to further investigate a causal role for sleep disruption in neurodegeneration in the human DS population provides an avenue for developing therapeutic strategies.

    We are aware of several limitations of our study due to experimental design. At the time of our experiments, the TcMAC21 mice available for prolonged sleep studies were few and included only male mice of a limited age range, described above. Expanding the sample size would provide for more statistical power to detect subtle sleep phenotypes, while the inclusion of females will be important to provide insight into the causes of the subtle sex and age differences DS patients, including males demonstrating increased N1 sleep26 as well as increased daytime sleepiness and napping behaviors compared to females.27 Finally, we also note that the average ages of our cohorts were approximately 1 week apart, with the EU mice being older than TS mice. One study comprehensively examining the influence of age on murine sleep-wake behavior during murine adolescence (postnatal day 15 through P87) did not find significant differences in total 24 hour sleep28 with age, making it less likely that age, itself was a confounding factor in our primary outcome. Similarly, NREM and REM sleep episode durations reached adult levels at P25 and P41, respectively, making it less likely that age governed the observed differences in sleep bout duration. Nevertheless, the impact of TcMAC21 on age-related changes in sleep would be of importance to investigate, first during adolescence given the subtle changes in NREM-REM balance that are seen during the first few months of development,28 as well as during longer time intervals (~12 months) during which there are gross changes in sleep architecture, including increased sleep during the phase.29

    Conclusion

    By recapitulating 93% of protein-coding genes from human chromosome 21q13, the TcMAC21 model represents one of the most accurate transgenic models of DS. Interestingly, in contrast to human DS sleep behavior, the TcMAC21 mice demonstrated significant sleep fragmentation, resulting in substantially decreased sleep during the dark phase, the primary wake time for mice. This model offers a unique platform for further investigate DS-related sleep disturbances, including sleep apnea, neuronal control of sleep, and long-term neurodevelopmental outcomes.

    Data Sharing Statement

    All data are available from the corresponding author upon reasonable request in accordance with journal guidelines.

    Acknowledgments

    We would like to acknowledge Khristine Amber Pasion and Zeynep Atak for their invaluable assistance in maintaining the TcMAC21 colony which was used for this project.

    Author Contributions

    JT and MS were responsible for data curation, formal analysis, and writing the original draft. TD and TH were responsible for study conceptualization, funding acquisition, supervision, and writing – reviewing/editing. All authors gave final approval of the version to be published; have agreed on the journal to which the article has been submitted; and agree to be accountable for all aspects of the work.

    Funding

    T.D. was supported by NINDS K08NS131529. T.H. was supported by R01NS116418 and R01NS136246.

    Disclosure

    All authors do not have any financial or non-financial relationships or conflicts of interest to disclose.

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    24. Duncan MJ, Guerriero LE, Kohler K, et al. Chronic fragmentation of the daily sleep-wake rhythm increases amyloid-beta levels and neuroinflammation in the 3xTg-AD mouse model of alzheimer’s disease. Neuroscience. 2022;481:111–122. doi:10.1016/j.neuroscience.2021.11.042

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  • Bird flu hiding in cheese? The surprising new discovery – Science Daily

    Bird flu hiding in cheese? The surprising new discovery – Science Daily

    1. Bird flu hiding in cheese? The surprising new discovery  Science Daily
    2. Bird flu virus survives in raw-milk cheese for months  News-Medical
    3. Live Avian Flu Can Survive in Raw Milk Cheese Up to Six Months  Food Poisoning Bulletin
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