ObjectiveTo systematically compare the differential regulation of the maternal-fetal interface cell lineages and communication networks in the CBA/J×DBA/2 mouse model of recurrent pregnancy loss （RPL） by the two classic therapeutic methods-tonifying the kidney to stabilize the fetus and invigorating the spleen to stabilize the fetus （Shoutai Wan， Juyuan Jian）-of traditional Chinese medicine （TCM） at the single-cell resolution and clarify their modern scientific connotations. MethodsFemale non-pregnant CBA/J mice were caged with male BALB/c （blank group） and DBA/2 （modeling group） mice separately. Pregnant mice in the modeling group were randomly grouped as follows： high/low-dose Shoutai Wan， high/low-dose Juyuan Jian， model （RPL）， and positive control （dydrogesterone）， with 10 mice in each group. Starting from the day after the detection of the vaginal plug， mice were administrated with drugs or an equal volume of normal saline by gavage for 10 consecutive days. After the intervention， the following indicators were measured. ① Macroscopic evaluation： general conditions， uterine wet weight， embryo loss rate， four coagulation parameters ［prothrombin time （PT）， activated partial thromboplastin time （APTT）， fibrinogen （FIB）， and thrombin time （TT）］， and peripheral blood estradiol （E₂） and progesterone （Pg） levels. The decidua with embryos was stained with hematoxylin-eosin （HE） and evaluated by transmission electron microscopy （TEM）. The expression of B-cell lymphoma-2 （Bcl-2）， vascular endothelial growth factor （VEGF）， angiotensin Ⅱ （AngⅡ）， matrix metalloproteinase-2 （MMP-2）， interleukin-6 （IL-6）， leukemia inhibitory factor （LIF）， CXC chemokine ligand 12 （CXCL12）， and microtubule-associated protein 1 light chain 3 homolog （LC3）Ⅰ/Ⅱ was quantified by Western blot. ② Mechanism analysis at the single-cell level： The decidua with embryos from the blank， model， high-dose Shoutai Wan， and high-dose Juyuan Jian groups （6 mice per group， with 3 single-cell samples per group， totaling 24 mice） were analyzed by the BD Rhapsody™ platform， and the whole-cell atlas was drawn by uniform manifold approximation and projection （UMAP） dimensionality reduction clustering combined with the single-cell mouse cell atlas （scMCA）. The differentially expressed genes （DEGs） and cell interaction networks were analyzed via Gene Ontology （GO）， Kyoto Encyclopedia of Genes and Genomes （KEGG）， and CellChat， and the protein-protein interaction （PPI） map of subtype cells was constructed. The CytoTRACE pseudo-temporal analysis was performed to explore the developmental trajectories of core immune cells （natural killer cells， NK cells） from maternal and fetal sources. Results① Pathological and Western blot results indicated that compared with the blank group， the RPL group showed an increase in the embryo loss rate （P<0.01）， down-regulated expression of Bcl-2， LIF， MMP-2， and Vegf in the decidua with embryos （P<0.05）， up-regulated protein levels of CXCL-12， AngⅡ， and IL-6 （P<0.05）， blocked angiogenesis， apoptosis-inflammation imbalance， and coagulation dysfunction. Both prescriptions dose-dependently reduced the abortion rate and restored the angiogenesis-inflammation balance， and Shoutai pill showed superior performance in restoring the E₂ level to the Pg level （P<0.05）. ② Single-cell transcriptome analysis indicated that compared with the blank group， the RPL group showed differences in multiple key cell populations such as decidual cells， trophoblast cells， endothelial cells， erythroblasts， NK cells， and macrophages at the maternal-fetal interface. Immunity and angiogenesis were the key links in RPL. Compared with the RPL group， high-dose Shoutai Wan reversed the changes of NK cells in the embryonic layer （upregulating the mRNA levels of 17 genes and downregulating the mRNA levels of 29 genes） and macrophages （upregulating the mRNA levels of 117 genes and downregulating the mRNA levels of 53 genes） through the regulation of gene expression. High-dose Shoutai pill regulated the immune cells to affect unfolded proteins， cell adhesion， and programmed cell death， thereby promoting decidualization and angiogenesis and modulating embryo-membrane development. High-dose Juyuan Jian regulated the key subgroups of NK cells （up-regulating the mRNA levels of 9 genes and down-regulating the mRNA levels of 17 genes） and macrophages （up-regulating the mRNA levels of 110 genes and down-regulating the mRNA levels of 81 genes）， which affected decidual inflammation and apoptosis and intervened in glycolysis. ③ The pseudo-temporal analysis and communication network indicated that the communication frequency of the RPL group decreased. High-dose Shoutai Wan restored maternal-fetal tolerance through pathways such as NKG2D， CDH5， GDF， and FASLG. High-dose Juyuan Jian enhanced the IL-6/LIFR/JAK/signal transducer and activator of transcription 3 （STAT3） and desmosome/SEMA6/tumor necrosis factor-like weak inducer of apoptosis （TWEAK） signaling to improve endometrial receptivity. The RPL group showed an increased proportion of toxic dNK7， a decreased proportion of reparative dNK4， and blocked embryo fNK1. High-dose Shoutai Wan down-regulated dNK7 and up-regulated dNK4. High-dose Juyuan Jian inhibited the terminal differentiation of dNK7 and up-regulated LILRB1， thus restoring the balance of cytotoxicity and repair. ConclusionBoth the kidney-tonifying and spleen-invigorating methods are effective in treating RPL. NK and macrophages are the key immune cells in the interaction between the embryo and the membrane. The kidney-tonifying method （Shoutai Wan） has an advantage in regulating the phenotypes of unfolded protein， cell adhesion， and programmed cell death， and shows expression characteristics closer to the physiological state in the regulation of NKG2D and CDH5 signals. The spleen-invigorating method （Juyuan Jian） has an advantage in regulating epithelial-mesenchymal transition （EMT）， angiogenesis， and glycolysis and shows higher communication intensity in the IL-6 and LIFR pathways.

[摘 要] 目的：构建负载STING激动剂DMXAA的锰卟啉金属有机框架纳米颗粒（DPM），探讨其对三阴性乳腺癌（TNBC）细胞4T1及其小鼠移植瘤的治疗效果。方法：通过物理吸附法制备 DPM 纳米颗粒，利用透射电镜、扫描电镜及纳米粒度电位仪表征其形貌与理化性质。常规培养4T1细胞，细胞实验分为对照组、超声辐照组（US组）、DPM治疗组（DPM组）和DPM治疗联合超声辐照组（DPM + US组），用CCK-8法检测细胞活性，免疫荧光法检测高迁移率族蛋白B1（HMGB1）和钙网蛋白（CRT）的表达，WB法检测STING通路相关蛋白的表达。构建4T1细胞移植瘤小鼠模型，分为四组，处理同细胞实验，测量肿瘤体积，免疫荧光法检测移植瘤组织中Ki-67、HMGB1、CRT和缺氧诱导因子-1ɑ（HIF-1ɑ）蛋白的表达，TUNEL法检测细胞凋亡，流式细胞术检测免疫细胞活化情况，对主要器官进行H-E染色，以评估纳米材料的体内安全性。结果：DPM呈梭形，平均粒径（268 ± 3.302）nm，电位（33.1 ± 0.87）mV。细胞实验中，DPM联合超声辐照可明显抑制4T1细胞的增殖（P < 0.001），提高4T1细胞中ROS水平（P < 0.001），诱导4T1细胞CRT表达上调（P < 0.001），HMGB1从细胞核中移至细胞质，激活STING信号通路[p-STING、p-TBK1、p-IRF3蛋白表达均显著增加（均P < 0.001）]。体内实验中，DPM联合超声辐照可显著抑制4T1细胞移植瘤生长（P < 0.001）并促进免疫细胞表型转化（P < 0.001），抑制移植瘤组织中Ki-67、HIF-1α蛋白表达（均P < 0.01），谷胱甘肽（GSH）产生（P < 0.01），促进CRT、HMGB1蛋白表达、ROS产生（P < 0.001），对主要器官结构无明显影响。结论： DPM联合超声辐照可通过激活STING通路显著抑制4T1细胞及其移植瘤的生长，诱导抗肿瘤免疫应答，且对主要器官无明显毒性。

Background: Adrenocortical hormones, particularly 11-oxygenated androgens, are pivotal in female reproductive health and fertility. Standardized detection kits and population-specific reference intervals are lacking in China, hindering related clinical applications. Methods: A HPLC-tandem mass spectrometry (HPLC-MS/MS) pipeline was developed, rigorously validated, and applied to simultaneously quantify corticosterone, cortisone, cortisol, 18-OH cortisol, androstenedione (A4), 11β-hydroxyandrostenedione (11-OH A4), dehydroepiandrosterone, and dehydroepiandrosterone sulfate in serum samples from 455 reproductive-aged women (18–45 yrs) in Guangxi, China. Age-dependent concentration trends were analyzed, and reference intervals stratified by age (2.5th to 97.5th percentiles) were established. Correlations with body-composition metrics, ethnicity, and the menstrual cycle were investigated. Results: The HPLC-MS/MS method demonstrated high precision (intra- and inter-assay CVs < 15%), accuracy, and sensitivity. All eight hormones exhibited significant age-related declines (P < 0.001 for seven hormones; P = 0.001 for 11-OH A4). Notably, 11-OH A4 levels were significantly lower in the 35–45-yr (3.05 nmol/L) and 25–34-yr (3.09 nmol/L) age groups than in the 18–24-yr (3.57 nmol/L) age group, whereas no significant difference was observed between the 35–45-yr and 25–34-yr age groups. Weak negative correlations were observed between the body mass index and corticosterone and cortisone levels, whereas ethnicity and the menstrual cycle showed no significant associations with hormone levels. Conclusions We developed an HPLC-MS/MS-based method for simultaneously quantifying eight adrenocortical hormones, including 11-OH A4, and defined age-specific reference intervals for reproductive-aged Chinese women. These findings advance the clinical utility of adrenocortical hormones in diagnosing and managing reproductive disorders.

Objective: To analyze the association between intraoperative blood transfusion and adverse perinatal outcomes(a composite of maternal death, hepatic encephalopathy, etc) in patients with acute fatty liver of pregnancy (AFLP), identify independent risk factors for these outcomes, and develop and validate a risk prediction model for early identification of high-risk patients. Methods: Clinical data of 56 AFLP patients admitted to our hospital from January 2008 to January 2025 were retrospectively analyzed. The chi-square test was used to compare the incidence of adverse perinatal outcomes between the transfusion group and the non-transfusion group. Univariate and multivariate Firth-penalized logistic regression analyses were performed to identify independent predictors of adverse outcomes. Based on the identified independent risk factors, a combined prediction model was developed. Internal validation was performed using the Bootstrap method (1 000 resamplings) to assess the model′s generalizability. Model performance was evaluated using receiver operating characteristic(ROC) curves and calibration curves. Finally, a nomogram for predicting adverse perinatal outcomes in AFLP was constructed. Results: Overall, 57.1%(32/56) of the patients received blood product transfusion during the termination of pregnancy. Multivariate Firth-penalized logistic regression analysis showed that, given the limited sample size, intraoperative blood transfusion had no independent statistical effect on adverse outcomes (OR=0.812, 95%CI:0.133-3.698, P=0.797). Mediation analysis also revealed no significant indirect effect. The independent risk factors were decreased preoperative fibrinogen (OR=14.062, 95%CI:2.389-126.656, P=0.003), pregnancy with infection (OR= 4.536, 95%CI:1.143-22.107, P=0.031), and cesarean section (OR=8.691, 95%CI:1.321-90.081, P=0.023). The combined prediction model achieved an AUC of 0.881 (95% CI:0.793-0.969, P<0.001), indicating good discrimination. At the optimal cut-off value, the sensitivity was 65.6%, specificity 95.8%, positive predictive value 95.5%, and negative predictive value 67.6%. Internal validation by the Bootstrap method showed that the predictive model had good discrimination and no obvious overfitting. The calibration curve demonstrated that the model′s predicted risk was consistent with the actual observed risk. Based on this, an AFLP perinatal adverse outcome nomogram model was constructed. Conclusion: Under the limited sample size of this study, no independent statistical effect of intraoperative blood product infusion on adverse perinatal outcomes in AFLP patients was detected. A multivariate Firth-penalized logistic regression model incorporating decreased preoperative fibrinogen, pregnancy with infection, and cesarean delivery demonstrated good discrimination and calibration. The derived nomogram may serve as an exploratory tool for early risk stratification and proactive intervention.

ObjectiveThis paper aims to investigate the traditional Chinese medicine syndrome types in patients with chronic hepatitis B （CHB） complicated by metabolic dysfunction-associated fatty liver disease （MAFLD） and explore the correlations between these syndrome types and clinical indicators， as well as ocular manifestation characteristics， thereby providing a reference for syndrome differentiation and treatment strategies in traditional Chinese medicine. MethodsGeneral data， information from the four diagnostic methods of traditional Chinese medicine， clinical indicators， and ocular manifestation data were collected from 506 patients with CHB complicated by MAFLD enrolled at the Public Health Clinical Center of Chengdu between June 2024 and December 2024. Cluster analysis， principal component analysis， and complex network models were employed to identify the distribution patterns of traditional Chinese medicine syndromes. Correlations between different syndrome types and clinical indicators， as well as ocular manifestation characteristics， were further analyzed. ResultsThe predominant syndromes identified in patients with CHB complicated by MAFLD were dampness and heat accumulation （51.58%）， liver depression with spleen deficiency （31.62%）， blood stasis obstructing collaterals （8.89%）， and Qi-Yin deficiency （7.91%）. No statistically significant differences were found among the four syndrome types in routine blood tests and liver function indicators. However， patients with the dampness and heat accumulation type exhibited significantly higher levels of total cholesterol （TC）， triglycerides （TG）， low-density lipoprotein cholesterol （LDL-C）， liver stiffness measurement （LSM）， controlled attenuation parameter （CAP）， and alpha-fetoprotein （AFP）， along with lower levels of high-density lipoprotein cholesterol （HDL-C）， compared with those with other syndrome types. Regarding ocular manifestations， the incidence of moon halo signs was significantly higher in patients with the blood stasis obstructing collaterals type than in those with other syndrome types. Additionally， the incidence in scleral zone 3 （corresponding to the large intestine） was higher in patients with the damp and heat accumulation type. ConclusionDampness and heat accumulation is the core syndrome type in patients with CHB complicated by MAFLD， commonly accompanied by spleen deficiency， liver depression， blood stasis， and Yin deficiency. A complex syndrome pattern characterized by a predominance of dampness and heat， along with a mixture of deficiency and excess， is formed. Different traditional Chinese medicine syndrome types are associated with distinct clinical indicators and ocular manifestation characteristics. Among them， patients with the dampness and heat accumulation type exhibit more pronounced metabolic disturbances and liver injury， whereas those with the blood stasis type show a higher incidence of moon halo signs. Abnormalities in scleral zone 3 are also more prevalent in patients with dampness and heat type.

ObjectiveTo investigate whether Bushen Huoxue prescription improves embryo implantation through regulating exosomal miRNA to enhance maternal-fetal interface communication based on Bushen Huoxue therapy. MethodsIn the animal experiment， all the rats （except for the blank group） were administered hydroxyurea （450 mg·kg^-1） via gavage for 10 d， as well as epinephrine （0.3 mg·kg^-1） and mifepristone （5.5 mg·kg^-1） via subcutaneous injection for 7 d to establish an implantation disorder model of kidney deficiency and blood stasis type. The Bushen Huoxue prescription （BSHX） groups were administered the prescription at different doses （7.30 g·kg^-1 for the high-dose group， 3.65 g·kg^-1 for the medium-dose group， and 1.83 g·kg^-1 for the low-dose group） via gavage. The dydrogesterone group was administered the corresponding medicine （2.63 mg·kg^-1） via gavage. After intervention for 10 days， uterine histopathological changes were observed via hematoxylin-eosin （HE） staining. Mucin （MUC1）， forkhead box protein O1 （FoxO1）， and homeobox A10 （HoxA10） expression levels were detected by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot. Cell experiment selected primary endometrial epithelial cells （EEC） and trophoblast cells （TC） as research subjects. Exosome-free medicated serum was prepared by ultracentrifugation and cultured in complete medium. Exosomes were isolated from cell supernatants by ultracentrifugation for cross-co-culture. After 48 h， migration and invasion abilities were assessed by scratch and Transwell assays. Sequencing was then performed on EEC-origin exosomal miRNA. ResultsThe model rats exhibited thin endometrium， along with reduced blood vessels， glandules， and pinopode numbers. BSHX improved endometrial morphology and increased pinopode numbers. MUC1， FoxO1， and HoxA10 expressions were downregulated in the model rats， while these parameters were upregulated after BSHX medium- and high-dose intervention. In the cell experiment， after exosome-free medicated serum intervention for 24 h， migration and invasion abilities were enhanced in the BSHX groups （P<0.01）. In EEC-origin exosomal miRNA sequencing， gene ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） analyses revealed enrichment in biological processes （gastrulation， neuronal differentiation， alongside cell development and regeneration）， involving the mitogen-activated protein kinase （MAPK）， FoxO1， Wnt， mammalian target of rapamycin （mTOR）， and tumor necrosis factor （TNF） signaling pathways. ConclusionBSHX promotes embryo implantation by improving endometrial receptivity via regulating exosomal miRNA. These findings provide potential targets for exosomal miRNA-based assisted reproductive strategies and a novel theoretical basis for infertility treatment by traditional Chinese medicine.

Objective To analyze the core elements of standardized diagnosis of occupational carpal tunnel syndrome (OCTS). Methods Data of first reported OCTS case in Guangdong Province were retrospectively analyzed, including occupational exposure history, clinical manifestations, auxiliary examinations, and on-site occupational health investigation results. Furthermore, standardized diagnostic approaches for such occupational diseases were explored. Results The patient worked in a garment manufacturing enterprise, engaged in fabric cutting. The patient had a occupational history of at least three consecutive years of repetitive right wrist work, holding a 15.2 kg electric cutting tool for a cumulative 6.6 hours per day, with awkward wrist movements repeated 13 times per minute. Clinical symptoms included numbness and reduced muscle strength in the areas of median nerve distribution of the right hand, which were aggravated during work and relieved after rest. Physical examination showed inability of right hand to perform fine motor opposition movements of the thumb with the index, middle, and ring fingers; decreased tactile, pain, vibration, graphesthesia, and kinesthetic sensations below the right wrist; and positive Tinel′s sign over the median nerve at the wrist crease level, Phalen′s test, and reverse Phalen′s test of right wrist. Both wrist high-frequency color Doppler ultrasonography and nerve conduction studies and electromyography indicated damage to the right median nerve. However, only the findings from high-frequency color Doppler ultrasonography of the wrist meet the diagnostic criteria specified in GBZ 336-2025 Diagnostic Standard for Occupational Carpal Tunnel Syndrome. On-site occupational health investigation confirmed that the work involved typical repetitive wrist work, and three co-workers in similar positions exhibited comparable symptoms. The case was ultimately diagnosed as OCTS. Conclusion The definitive diagnosis of OCTS requires comprehensive evaluation of four core elements: a clear occupational exposure history, typical clinical symptoms and signs, characteristic objective examination findings, and quantitative on-site occupational health investigation data. Only when these four elements corroborate each other and non-occupational causative factors are excluded can a definitive diagnosis be made.

Objective: To analyze the characteristics and key risk factors of abnormal blood disposal throughout the 2024-2025 blood collection and supply process based on nationwide multi-site monitoring data, develop a comprehensive prevention and control strategy across the entire chain, reduce preventable blood waste, and ensure clinical blood safety. Methods: According to the Guidelines for Haemovigilance (T/CSBT 001-2026), we extracted 74 cases of abnormal blood disposal events reported by the China Blood Transfusion Association′s Blood Safety Monitoring System from January 1 2024, to December 31 2025, and conducted descriptive statistical analysis on the distribution of events, types of deviations, root causes, and classifications of disposal manifestations. Results: A total of 664 adverse events related to blood collection and supply were reported over two years, with 74 cases involving discarded abnormal blood units, accounting for 11.1% of the total. Among these, blood component preparation (40.5%) and blood collection (37.8%) remained traditional high-risk stages. The proportion of blood waste occurring during storage, distribution, and transportation increased from 7.7% in 2024 to 34.3% in 2025. Deviations from standard operating procedures (SOP) by personnel were the primary cause, accounting for 74.3% of cases. Discarded blood units were categorized into four types: compromised closed blood bag systems (40 cases), mainly due to inadequate heat sealing, physical damage, or dropped bags; blood out of the cold chain (12 cases), primarily resulting from blood being left at work sites; imbalanced proportion of preservation solution (10 cases), mostly caused by blood collection volumes exceeding the standard capacity of blood bags; and other causes (12 cases). Conclusion: Human operational errors are the core cause of abnormal blood waste. The collection and preparation stages have long been at high-risk, and the risks in the storage and transportation stages have been increasing year by year. We should focus on standardizing personnel management, improving specialized operation norms such as heat sealing, transportation, and storage, establishing a full-process intelligent cold chain monitoring and multi-node visual verification mechanism, and systematically reducing the occurrence rate of abnormal blood waste through a closed-loop quality intervention system, thereby improving the utilization efficiency of voluntary blood donation resources.

ObjectiveThis study aimed to investigate the mechanism by which Luoshi Neiyi prescription treats endometriosis （EMs） through regulating ferroptosis， and to screen key metabolites and analyze their association with ferroptosis. MethodsClinical samples of normal endometrium from patients without EMs and eutopic and ectopic endometrium from EMs patients （10 cases each） were collected and divided into control group， eutopic group， and EMs group. Hematoxylin-eosin （HE） staining was performed to observe ectopic lesions of EMs. Immunohistochemistry was used to detect the expression of solute carrier family 7 member 11 （SLC7A11） and glutathione peroxidase 4 （GPX4）. Enzyme-linked immunosorbent assay （ELISA） was adopted to determine the levels of malondialdehyde （MDA）， ferrous ion （Fe²⁺）， GPX4 and glutathione （GSH） in endometrial tissues， as well as serum levels of Fe²⁺， GPX4 and GSH. Real-time quantitative polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of SLC7A11， GPX4， transferrin receptor （TFR） and ferritin heavy chain 1 （FTH1）. In the in vitro experiment， primary stromal cells were isolated from ectopic lesions of EMs patients. Cell counting kit-8 （CCK-8） was used to determine the optimal concentration of drug-containing serum for intervention. The level of reactive oxygen species （ROS） was measured， and Real-time PCR was applied to detect ferroptosis-related indicators. In the animal experiments， an EM rat model was established， and the rats were randomly assigned to the sham operation group， EMs group， low-dose Luoshi Neiyi Formula group （7.87 g·kg^-1）， high-dose Luoshi Neiyi prescription group （15.74 g·kg^-1）， and danazol group （42 mg·kg^-1）. Untargeted metabolomics detection and pathway enrichment analysis were conducted on serum samples from patients and rats. Spearman correlation analysis was performed to assess the relationship between differential metabolites and ferroptosis indicators. The correlations between differential metabolites in patient endometrium and serum and key ferroptosis indicators （GPX4， Fe²⁺， MDA， GSH） as well as ferroptosis-related mRNAs （GPX4， SLC7A11， FTH1） were analyzed， and correlation heatmaps were generated accordingly. ResultsCompared with normal eutopic endometrium， ectopic lesions in EMs patients showed glandular disorganization and stromal fibrosis. In ectopic endometrium， the contents of MDA， ROS， and Fe²⁺ decreased， while GPX4 level increased， and the mRNA expression of SLC7A11 and GPX4 was upregulated （P<0.05， P<0.01）. In serum， the levels of GPX4 and Fe²⁺ were elevated， whereas the GSH level declined， suggesting abnormalities in ferroptosis-related pathways in ectopic lesions （P<0.05， P<0.01）. After intervention with Luoshi Neiyi prescription-containing serum， the intracellular ROS level in ectopic endometrial stromal cells was elevated， the mRNA expression of SLC7A11 and GPX4 was downregulated， and TFR mRNA expression was upregulated （P<0.05， P<0.01）. Metabolomics analysis revealed 1104 and 198 differential metabolites in EMs patients and EMs rats， respectively， compared with their corresponding control groups， and both low-dose and high-dose Luoshi Neiyi prescription were found to regulate this metabolic disturbance， with the core regulatory pathways mainly involving arginine and proline metabolism. Correlation analysis showed that the glycerophospholipids including PI（16∶0/17∶0） and PI［18∶2（9Z，12Z）］ were negatively correlated with GPX4 and positively correlated with MDA， while 17α-hydroxyprogesterone was positively correlated with GPX4， SLC7A11， and FTH1 q<0.05）. ConclusionLuoshi Neiyi prescription may systematically ameliorate disease-associated metabolic dysregulation via modulation of the serum arginine and proline metabolism pathway， and may regulate ferroptosis in ectopic lesions through a mechanism potentially linked to the serum glycerophospholipid and steroid metabolism pathways. Collectively， these findings provide experimental evidence for the clinical application of Luoshi Neiyi prescription.

ObjectiveThis study aimed to investigate the mechanism by which Luoshi Neiyi prescription treats endometriosis （EMs） through regulating ferroptosis， and to screen key metabolites and analyze their association with ferroptosis. MethodsClinical samples of normal endometrium from patients without EMs and eutopic and ectopic endometrium from EMs patients （10 cases each） were collected and divided into control group， eutopic group， and EMs group. Hematoxylin-eosin （HE） staining was performed to observe ectopic lesions of EMs. Immunohistochemistry was used to detect the expression of solute carrier family 7 member 11 （SLC7A11） and glutathione peroxidase 4 （GPX4）. Enzyme-linked immunosorbent assay （ELISA） was adopted to determine the levels of malondialdehyde （MDA）， ferrous ion （Fe²⁺）， GPX4 and glutathione （GSH） in endometrial tissues， as well as serum levels of Fe²⁺， GPX4 and GSH. Real-time quantitative polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of SLC7A11， GPX4， transferrin receptor （TFR） and ferritin heavy chain 1 （FTH1）. In the in vitro experiment， primary stromal cells were isolated from ectopic lesions of EMs patients. Cell counting kit-8 （CCK-8） was used to determine the optimal concentration of drug-containing serum for intervention. The level of reactive oxygen species （ROS） was measured， and Real-time PCR was applied to detect ferroptosis-related indicators. In the animal experiments， an EM rat model was established， and the rats were randomly assigned to the sham operation group， EMs group， low-dose Luoshi Neiyi Formula group （7.87 g·kg^-1）， high-dose Luoshi Neiyi prescription group （15.74 g·kg^-1）， and danazol group （42 mg·kg^-1）. Untargeted metabolomics detection and pathway enrichment analysis were conducted on serum samples from patients and rats. Spearman correlation analysis was performed to assess the relationship between differential metabolites and ferroptosis indicators. The correlations between differential metabolites in patient endometrium and serum and key ferroptosis indicators （GPX4， Fe²⁺， MDA， GSH） as well as ferroptosis-related mRNAs （GPX4， SLC7A11， FTH1） were analyzed， and correlation heatmaps were generated accordingly. ResultsCompared with normal eutopic endometrium， ectopic lesions in EMs patients showed glandular disorganization and stromal fibrosis. In ectopic endometrium， the contents of MDA， ROS， and Fe²⁺ decreased， while GPX4 level increased， and the mRNA expression of SLC7A11 and GPX4 was upregulated （P<0.05， P<0.01）. In serum， the levels of GPX4 and Fe²⁺ were elevated， whereas the GSH level declined， suggesting abnormalities in ferroptosis-related pathways in ectopic lesions （P<0.05， P<0.01）. After intervention with Luoshi Neiyi prescription-containing serum， the intracellular ROS level in ectopic endometrial stromal cells was elevated， the mRNA expression of SLC7A11 and GPX4 was downregulated， and TFR mRNA expression was upregulated （P<0.05， P<0.01）. Metabolomics analysis revealed 1104 and 198 differential metabolites in EMs patients and EMs rats， respectively， compared with their corresponding control groups， and both low-dose and high-dose Luoshi Neiyi prescription were found to regulate this metabolic disturbance， with the core regulatory pathways mainly involving arginine and proline metabolism. Correlation analysis showed that the glycerophospholipids including PI（16∶0/17∶0） and PI［18∶2（9Z，12Z）］ were negatively correlated with GPX4 and positively correlated with MDA， while 17α-hydroxyprogesterone was positively correlated with GPX4， SLC7A11， and FTH1 q<0.05）. ConclusionLuoshi Neiyi prescription may systematically ameliorate disease-associated metabolic dysregulation via modulation of the serum arginine and proline metabolism pathway， and may regulate ferroptosis in ectopic lesions through a mechanism potentially linked to the serum glycerophospholipid and steroid metabolism pathways. Collectively， these findings provide experimental evidence for the clinical application of Luoshi Neiyi prescription.