1.Exploration on Mechanism of Topical Treatment of Allergic Contact Dermatitis in Mice with Portulacae Herba Based on Nrf2/HO-1/NF-κB Signaling Pathway
Xiaoxue WANG ; Guanwei FAN ; Xiang PU ; Zhongzhao ZHANG ; Xia CHEN ; Ying TANG ; Nana WU ; Jiangli LUO ; Xiangyan KONG
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(3):115-123
ObjectiveTo investigate the mechanism of topical treatment of allergic contact dermatitis (ACD) mice with Portulacae Herba based on the nuclear factor E2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1)/nuclear factor-κB (NF-κB) signaling pathway. MethodsA total of 70 6-week-old specific pathogen free (SPF) female Kunming mice were adaptively fed for 1 week and randomly divided into blank group, model group, compound dexamethasone acetate cream group (2.075×10-2 g·g-1), blank matrix cream group, low-dose Portulacae Herba cream group (0.1 g·g-1), high-dose Portulacae Herba cream group (0.2 g·g-1), and Portulacae Herba + inhibitor group (0.2 g·g-1 + 30 mg·kg-1 ML385), with 10 mice in each group. One day before the experiment, the mice were shaved on the neck and back. Except for the blank group, the mice in the other groups were treated with 2,4-dinitrochlorobenzene (DNCB) to establish an ACD model. After respective administration, the skin lesion of the mice was scored, and the histopathological changes of the skin were stained with hematoxylin-eosin (HE). Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of interleukin-6 (IL-6), interleukin-1β (IL-1β), reactive oxygen species (ROS), superoxide dismutase (SOD) activity, and malondialdehyde (MDA) in serum of mice. The expression of Nrf2/HO-1/NF-κB signaling pathway-related proteins in mouse skin tissue was detected by immunohistochemistry (IHC), Western blot, and real-time fluorescence quantitative polymerase chain reaction (Real-time PCR). ResultsCompared with the blank group, the mice in the model group had an increased skin lesion score (P<0.01), severe pathological damage to skin tissue, increased content of IL-1β, IL-6, ROS, and MDA in their serum (P<0.01), and decreased content of SOD (P<0.01). In addition, the mRNA and protein expression levels of Nrf2, HO-1, and nuclear factor-κB inhibitor α (IκBα) in skin tissue were up-regulated (P<0.01), while the protein expression levels of phosphorylated (p)-IκBα and p-NF-κB p65 and the mRNA expression of NF-κB p65 were down-regulated (P<0.01). Compared with the model group and the blank matrix cream group, the mice treated with Portulacae Herba had a decreased skin lesion score (P<0.01), reduced pathological damage to skin tissue, decreased content of IL-1β, IL-6, ROS, and MDA in their serum (P<0.01), and increased content of SOD (P<0.01). Additionally, the mRNA and protein expression levels of Nrf2, HO-1, and IκBα in skin tissue were down-regulated (P<0.05,P<0.01), and the protein expression levels of p-IκBα and p-NF-κB p65 and the mRNA expression of NF-κB p65 were up-regulated (P<0.05,P<0.01). Compared with the Portulacae Herba + inhibitor group, the high-dose Portulacae Herba cream group had a decreased skin lesion score (P<0.01), alleviated pathological damage to skin tissue, decreased content of IL-1β, IL-6, ROS, and MDA in the serum of mice (P<0.05,P<0.01), and increased content of SOD (P<0.01). The protein expression levels of Nrf2, HO-1, and IκBα and the mRNA expression of Nrf2 and HO-1 in skin tissue were up-regulated (P<0.05,P<0.01), and the protein expression levels of p-IκBα and p-NF-κB p65 and the mRNA expression of NF-κB p65 were down-regulated (P<0.05). ConclusionPortulacae Herba can improve DNCB-induced ACD skin damage in mice by regulating the Nrf2/HO-1/NF-κB signaling pathway.
2.Analysis of Application of Animal Model of Spleen Deficiency and Dampness Syndrome Based on Data Mining
Qingqian YU ; Yifei ZHANG ; Zehan ZHANG ; Weiyue ZHANG ; Yuebo WANG ; Fengzhi WU ; Feng LI
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(3):235-243
ObjectiveThe research focuses on developing modeling and evaluation methodologies for an animal model exhibiting spleen deficiency and dampness excess syndrome, with the aim of standardizing such animal models for future reference. MethodsBy conducting a literature search on animal models of spleen deficiency and dampness excess syndrome, relevant publications meeting inclusion and exclusion criteria will be identified based on publication date, data source, types of diseases involved, animal characteristics, modeling methods, modeling duration, macroscopic syndrome assessment indicators, macroscopic quantification indicators, laboratory testing parameters, intervention approaches, positive controls and application context. A database will be established to facilitate the extraction of this information for quantitative analysis, statistical evaluation, and visual representation. ResultsA total of 137 literature articles meeting the standards have been included in the research. The primary animal species used in animal models of spleen deficiency and dampness excess are SD rats. Modeling methods include single-factor, dual-factor composite, and triple-factor composite methods, with various models widely applied in validation of pharmacological effects and mechanistic explorations. Evaluation indices of animal models for spleen deficiency and dampness excess primarily consist of macroscopic syndrome evaluation indicators and macroscopic quantitative indicators. Laboratory testing indicators are mostly related to research areas such as fluid metabolism and gastrointestinal function. The most commonly studied herbal formulas currently include Shenling Baizhu San and Pingwei San, with natural recovery and the use of the western medicine metronidazole as the most frequently used positive controls. ConclusionThe application of animal models for spleen deficiency and dampness excess is gradually increasing, with various modeling methods already simulating the typical characteristics of this syndrome pattern. However, there are still many areas that are worth contemplating and improving. This study aims to provide reference and ideas for the standardization of symptom names in animal models of spleen deficiency and dampness excess, as well as for the improvement of model construction and evaluation systems.
3.The Effect of Modified Ditan Decoction (涤痰汤) on Cognitive Function and Resting-State Functional Magnetic Resonance Imaging of the Brain in Chronic Intermittent Hypoxia Model Rats
Naijie CHEN ; Xiaoting WANG ; Fengsheng XU ; Shuanghong SHEN ; Zuanfang LI ; Qin CHEN ; Jin CHEN ; Runhua WU
Journal of Traditional Chinese Medicine 2025;66(1):71-78
ObjectiveTo explore the effect of modified Ditan Decoction (涤痰汤) on chronic intermittent hypoxia cognitive function and the potential function mechanism. MethodsTwenty-four Sprague-Dawley (SD) rats were randomly divided into a normal group, a model group, and a modified Ditan Decoction group, with eight rats in each group. Rats in the modified Ditan Decoction group were administered the decoction by gavage at 14.8 ml/(kg·d), while the normal group and the model group received the same dose of normal saline. Thirty minutes after daily gavage, the rats in all three groups were placed in an intermittent hypoxia chamber. The oxygen concentration for the model group and the modified Ditan Decoction group was adjusted daily for 8 hours using a computer program to establish the model, while the normal group was exposed to the same airflow rate of ambient air. The intervention was continued for 12 weeks to establish a chronic intermittent hypoxia rat model. The Y-maze test was used to evaluate spatial working memory in the rats. Resting-state functional magnetic resonance imaging (rs-fMRI) was performed to detect whole-brain regional homogeneity (ReHo) and seed-based functional connectivity (FC). Brain regions showing significant differences in rs-fMRI were selected for further analysis. Immunofluorescence was used to detect β-amyloid (Aβ) deposition and the number of ionized calcium-binding adapter molecule 1 (IBA1)-positive microglial cells. Immunohistochemistry was employed to assess the expression of synaptophysin (SYP), the excitatory synapse marker vesicular glutamate transporter 1 (Vglut1), and the inhibitory synapse marker vesicular γ-aminobutyric acid transporter (VGAT). ResultsCompared with the normal group, the model group showed a reduced spontaneous alternation rate in the Y-maze test. The smoothed Z-score standardized regional homogeneity (SzReHo) value in the left entorhinal cortex significantly increased, and the FC value from this seed point to the left basal forebrain significantly reduced. Additionally, the model group exhibited significantly higher Aβ fluorescence intensity and Iba1 positivity in the left entorhinal cortex, decreased expression of SYP, Vglut1, and VGAT, along with an increased Vglut1/VGAT ratio (P<0.05 or P<0.01). Compared to the model group, the modified Ditan Decoction group demonstrated an increased spontaneous alternation rate, a significantly reduced SzReHo value in the left entorhinal cortex, and a significantly increased FC value from this region to the left basal forebrain. Furthermore, this group showed significantly lower Aβ fluorescence intensity and Iba1 positivity in the left entorhinal cortex, increased levels of SYP, Vglut1, and VGAT, and a decreased Vglut1/VGAT ratio (P<0.05 or P<0.01). ConclusionModified Ditan Decoction can reconstruct the projection from the left basal forebrain to the entorhinal cortex in chronic intermittent hypoxia, thereby reducing Aβ aggregation and excessive microglial activation in the left entorhinal cortex. This process improves the excitation/inhibition imbalance caused by synaptic remodeling, ultimately enhancing cognitive function in rats of chronic intermittent hypoxia.
4.Exploration of the Application of Fengfu (GV 16) Acupoint in BIAN Que Heart Book (《扁鹊心书》)
Yawei ZHAO ; Haoying LI ; Lintong WEN ; Hefei WANG ; Wei WANG ; Hongyu WU ; Shijiang SUN
Journal of Traditional Chinese Medicine 2025;66(1):98-101
By examining the records related to the Fengfu (GV 16) acupoint in BIAN Que Heart Book (《扁鹊心书》) compiled by the Song Dynasty physician DOU Cai, this study analyzed various aspects, including the differentiation of conditions treated with Fengfu (GV 16) acupoint, the theoretical foundation for selection of Fengfu (GV 16) acupoint, the application of needling manipulation, and the sensation of obtaining qi during acupuncture. The findings suggest that DOU Cai's approach to utilizing Fengfu (GV 16) acupoint differs from traditional methods, particularly emphasizing the effectiveness of achieving a sensation of heat and numbness. His unique techniques include transverse insertion at Fengfu (GV 16) acupoint and penetrated insertion to Fengchi (GB 20) and Yifeng (TE 17) acupoints. The records of Fengfu (GV 16) acupoint in BIAN Que Heart Book provide a valuable reference for its modern clinical application and further development.
5.Effects of bioactive peptides combined with probiotics on serum uric acid in patients with hyperuricemia
HAN Dan ; ZHAO Ya ; HUANG Enshan ; YE Shuhua ; WANG Wanjin ; WU Fangmin ; WANG Dingliang ; ZHANG Ronghua
Journal of Preventive Medicine 2025;37(1):40-45
Objective:
To evaluate the effect of bioactive peptides combined with probiotics on serum uric acid (SUA) in patients with hyperuricemia (HUA), so as to provide the evidence for prevention and treatment of HUA.
Methods:
The patients with HUA aged 18 to 65 years were selected and randomly divided into an intervention group and a control group. The patients in the intervention group received bioactive peptides combined with probiotics for 28 days at a dose of 3 g/d, while the patients in the control group received an equal dose of placebos. Demographic information, body mass index (BMI), blood pressure and blood lipid were collected through questionnaire surveys, physical examination and laboratory tests. SUA levels were detected before and after 14 days and 28 days of interventions. The differences of SUA levels between the two groups were compared using generalized estimation equation.
Results:
Totally 108 patients with HUA were recruited, including 54 patients in the intervention group and 53 patients in the control group (1 dropout). Before interventions, there were no statistically significant differences in gender, age, course of HUA, exercise duration, frequency of alcohol consumption, frequency of meat broth consumption, BMI, prevalence of hypertension and prevalence of dyslipidemia between the two groups (all P>0.05). After 14 days of interventions, the SUA levels of the patients in the intervention group decreased by 3.00 μmol/L, while those in the control group increased by 7.00 μmol/L. After 28 days of interventions, the SUA levels of the patients in the intervention group and the control group decreased by 26.00 μmol/L and 16.00 μmol/L, respectively. However, there was no statistically significant interaction between the intervention time and group (both P>0.05). Subgroup analysis showed that after 28 days of interventions, the decrease in SUA levels in the patients aged 55 years and older and without hypertension in the intervention group was greater than those in the control group (both P<0.05).
Conclusions
Bioactive peptides combined with probiotics showed no significant difference in reducing SUA levels in patients with HUA compared to the control group. The effect was more significant for patients aged 55 years and older and without hypertension.
6.Pathogenic characteristics and drug sensitivity analysis of hospital-acquired infections in lung transplant recipients: a single-center 5-year retrospective study
Sangsang QIU ; Qinfen XU ; Bo WU ; Xiaojun CAI ; Qinhong HUANG ; Dapeng WANG ; Chunxiao HU ; Jingyu CHEN
Organ Transplantation 2025;16(1):114-121
Objective To analyze the characteristics of postoperative hospital-acquired infections and drug sensitivity in lung transplant recipients over the past 5 years in a single center. Methods A total of 724 lung transplant recipients at Wuxi People's Hospital from January 2019 to December 2023 were selected. Based on the principles of hospital-acquired infection diagnosis, a retrospective analysis was conducted on the hospital infection situation and infection sites of lung transplant recipients, and an analysis of the distribution of hospital-acquired infection pathogens and their antimicrobial susceptibility test status was performed. Results Among the 724 lung transplant recipients, 275 cases of hospital-acquired infection occurred, with an infection rate of 38.0%. The case-time infection rate decreased from 54.2% in 2019 to 22.8% in 2023, showing a downward trend year by year (Z=30.98, P<0.001). The main infection site was the lower respiratory tract, accounting for 73.6%. The pathogens were mainly Gram-negative bacteria, with the top four being Acinetobacter baumannii (37.1%), Pseudomonas aeruginosa (17.3%), Klebsiella pneumoniae (13.7%), and Stenotrophomonas maltophilia (13.4%), with imipenem resistance rates of 89%, 53%, 58% and 100%, respectively. Gram-positive bacteria were mainly Staphylococcus aureus (3.6%), with a methicillin resistance rate of 67%. Conclusions Over the past 5 years, the hospital-acquired infections in lung transplant recipients have shown a downward trend, mainly involving lower respiratory tract infections, with the main pathogens being Acinetobacter baumannii, Pseudomonas aeruginosa and Klebsiella pneumoniae, all of which have high resistance rates to imipenem.
7.A survival prediction model for kidney graft based on the kidney donor profile index, time-zero biopsy and donor’s age
Chengxi JIANG ; Shunliang YANG ; Xia GAO ; Liqian WU ; Jiashu LIU ; Dong WANG
Organ Transplantation 2025;16(1):122-130
Objective To construct a predictive model for the survival of transplant kidneys after kidney transplantation. Methods The clinical data of 366 kidney transplant recipients and donors were retrospectively analyzed, and the recipients were divided into low-risk group (n=101), medium-risk group (n=189), and high-risk group (n=76) based on the kidney donor profile index (KDPI). Each group was further divided into Remuzzi score ≤3 group and Remuzzi score >3 group based on time-zero biopsy Remuzzi scores. Kaplan-Meier method was used to analyze the survival of transplant kidneys. Univariate and multivariate Cox regression analyses were performed to identify risk factors affecting long-term survival after kidney transplantation. A predictive model for transplant kidney survival was established and a nomogram was drawn. The predictive performance of the model was evaluated using the receiver operating characteristic (ROC) curve and the area under the curve (AUC). Results The median KDPI was 65%, and the median Remuzzi score was 3. The 5-year survival rate of transplant kidneys was 83.5%. Kaplan-Meier survival curves showed that in the KDPI medium-risk and KDPI high-risk groups, the subgroup with lower Remuzzi score had a higher survival rates of transplant kidneys than the subgroup with higher Remuzzi score. Univariate and multivariate Cox regression analyses showed that KDPI, Remuzzi score, and donor’s age were independent risk factors for transplant kidney loss (all P<0.05). The ROC curve showed that the AUC of the nomogram prediction model established based on independent risk factors for the 1, 3 and 5-year survival rates of transplant kidneys were 0.91, 0.93 and 0.94 for the training set, and 0.89, 0.85 and 0.88 for the validation set. Calibration curves shows good consistency between the training and validation sets of the model. Conclusions The nomogram predictive model based on KDPI, time-zero biopsy Remuzzi score and donor’s age has good predictive value for transplant kidney survival.
8.Mechanism of Modified Shaofu Zhuyutang in Antagonising Ectopic Endometrial Tissue Fibrosis Based on Cellular Pyroptosis Mediated by TRL4/NF-κB/NLPR3 Signaling Pathway
Zuoliang ZHANG ; Jiaxing WANG ; Wanrun WANG ; Xiangyu LIN ; Bin YUE ; Zhirui ZHANG ; Yinan WANG ; Yaling YANG ; Dongqing WEI ; Cancan HUANG ; Quansheng WU
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(4):19-28
ObjectiveTo investigate the mechanism of action of modified Shaofu Zhuyutang in antagonizing cellular pyroptosis and fibrosis in ectopic endometrial tissues of endometriosis through the Toll-like receptor 4/nuclear factor-κB/NOD-like receptor protein 3 (TRL4/NF-κB/NLPR3) signaling pathway. MethodsSeventy-two SPF-grade female SD rats were randomly divided into a sham-operated group (n = 12) and a modeling group (n = 60). The rats in the sham-operated group underwent a caesarean section, while the rats in the modeling group were used to establish an endometriosis model through the auto-transplantation method. After successful modeling, the animals were randomly divided into the model group, progesterone group (0.25 mg·kg-1), and modified Shaofu Zhuyutang low-, medium-, and high-dose groups (7.5, 15, 30 g·kg-1), with 12 animals in each group. After 4 weeks of drug administration, voluntary activity and heat pain latency were observed. The rats were sacrificed for tissue collection, and Masson staining were used to observe histopathological changes in the endometrial tissues. Enzyme-linked immunosorbent assay (ELISA) was used to measure serum levels of interleukin-18 (IL-18), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and transforming growth factor-β (TGF-β). Immunohistochemistry (IHC) was used to detect the protein expression area of tumor necrosis factor-related factor 6 (TRAF6) and NLPR3 in the endometrial tissues. Immunofluorescence (IF) was used to detect the relative fluorescence intensity of Caspase-1 and gasdermin D (GSDMD) in the endometrial tissues. Western blot was employed to measure the relative expression of TRL4, myeloid differentiation factor 88 (MyD88), TRAF6, NF-κB p65, phosphorylated NF-κB p65 (p-NF-κB p65), and NLPR3 proteins in endometrial tissues. Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) was used to detect the mRNA expression of TRL4, MyD88, TRAF6, NF-κB, and NLPR3 in the endometrial tissues. ResultsCompared with the sham-operated group, rats in the model group showed reduced voluntary activity and shorter heat pain latency. Serum levels of IL-18, IL-1β, TNF-α, and TGF-β were elevated. The relative expression areas of TRAF6 and NLPR3 proteins were increased, and the relative fluorescence intensity of Caspase-1 and GSDMD was enhanced. The relative expression of TRL4, MyD88, TRAF6, NF-κB p65, p-NF-κB p65, and NLPR3 proteins, along with the expression of TRL4, MyD88, TRAF6, NF-κB, and NLPR3 mRNA, were significantly increased (P<0.01). Compared with the model group, rats in the progesterone group and the modified Shaofu Zhuyutang medium- and high-dose groups exhibited improved voluntary activity, longer heat pain latency, the fibrosis of endometrial tissue is alleviated. Serum levels of IL-18, IL-1β, TNF-α, and TGF-β were decreased. The relative expression areas of TRAF6 and NLPR3 proteins decreased, and the relative fluorescence intensity of Caspase-1 and GSDMD weakened. The relative expression of TRL4, MyD88, TRAF6, p-NF-κB p65, NLPR3 proteins, and TRL4, MyD88, TRAF6, NF-κB, and NLPR3 mRNA expression were reduced (P<0.05, P<0.01). ConclusionModified Shaofu Zhuyutang may play a therapeutic role in endometriosis by interfering with key proteins in the TRL4/NF-κB/NLPR3 signaling pathway, reducing NLRP3 inflammasome-induced cellular pyroptosis, antagonizing the fibrosis process in ectopic endometrial tissues, improving the inflammatory microenvironment in the pelvic cavity, and alleviating pain.
9.Mechanism of Modified Shaofu Zhuyutang in Treatment of Endometriosis Based on EGFR/PI3K/Akt Signaling Pathway
Yaling YANG ; Wanrun WANG ; Zuoliang ZHANG ; Xiangyu LIN ; Jiaxing WANG ; Cancan HUANG ; Xiujia JI ; Quansheng WU
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(4):29-38
ObjectiveTo observe the effects of modified Shaofu Zhuyutang on key proteins of the epidermal growth factor receptor (EGFR)/phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway in SD rats with endometriosis. MethodsAfter successful establishment of an endometriosis model in 60 female SD rats of SPF grade via the auto-transplantation method, the rats were randomly divided into a model group, modified Shaofu Zhuyutang high-, medium-, and low-dose groups, and a gestrinone group, with another 12 rats serving as a blank group. The blank and model groups were administered 10 mL·kg-1 normal saline, while the high-, medium-, and low-dose groups received 30, 15, and 7.5 g·kg-1 modified Shaofu Zhuyutang, respectively. The gestrinone group was administered 0.25 mg·kg-1 gestrinone suspension. After four weeks of treatment, uterine contractions were induced with 2 U of oxytocin, and the writhing response of rats was observed. After 24 h, the rats were euthanized, and the weight and volume of ectopic endometrial tissue were recorded. Hematoxylin-eosin (HE) staining was used to observe pathological changes in endometrial tissues, while the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay was used to evaluate the apoptosis rate of endometrial tissues. Immunofluorescence was used to detect the relative expression areas of the B-cell lymphoma-2 gene-associated promoter (Bad) and B-cell lymphoma-2 (Bcl-2) proteins in endometrial tissues. Serum levels of interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), epidermal growth factor (EGF), and EGFR were measured by enzyme-linked immunosorbent assay (ELISA). The relative protein expression levels of EGFR, PI3K, phosphorylated PI3K (p-PI3K), Akt, and phosphorylated Akt (p-Akt) in endometrial tissues were analyzed by Western blot. Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) was used to detect the mRNA expression levels of EGFR, PI3K, and Akt. ResultsCompared with the blank group, the model group showed endometrial thickening, glandular and mesenchymal hyperplasia, a significant decrease in the relative expression area of Bad in ectopic endometrial tissues, a significant increase in the relative expression area of Bcl-2, and a significant reduction in the apoptosis rate as indicated by TUNEL staining. Serum levels of IL-1β, IL-6, TNF-α, EGF, and EGFR were significantly elevated (P<0.01). The relative protein expression levels of EGFR, PI3K, p-PI3K, Akt, and p-Akt, as well as the mRNA expression levels of EGFR, PI3K, and Akt, were also significantly increased (P<0.01). Compared with the model group, the high- and medium-dose groups of modified Shaofu Zhuyutang and the gestrinone group exhibited reduced glandular and mesenchymal hyperplasia to varying degrees, with dilated glandular lumens. The number of writhing responses was significantly reduced, the latency to writhing response was significantly prolonged, and the weight and volume of ectopic endometrial tissue were significantly decreased. The relative expression area of Bad in ectopic endometrial tissue was significantly increased, the relative expression area of Bcl-2 was significantly decreased, and the apoptosis rate was significantly elevated as shown by TUNEL staining. Serum levels of IL-1β, IL-6, TNF-α, EGF, and EGFR were significantly reduced, and the relative protein expression levels of EGFR, PI3K, p-PI3K, Akt, and p-Akt, as well as the mRNA expression levels of EGFR, PI3K, and Akt, were significantly decreased (P<0.05,P<0.01). ConclusionModified Shaofu Zhuyutang may exert therapeutic effects on endometriosis by interfering with key proteins of the EGFR/PI3K/Akt signaling pathway and inducing apoptosis in ectopic endometrial tissue.
10.Effect of Modified Shaofu Zhuyutang on Ferroptosis in Ectopic Endometrial Tissues of Rats with Endometriosis Based on MDM4/p53/GPX4 Signaling Pathway
Zuoliang ZHANG ; Xiangyu LIN ; Wanrun WANG ; Jiaxing WANG ; Yaling YANG ; Quansheng WU
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(4):39-47
ObjectiveTo investigate the mechanism of modified Shaofu Zhuyutang in inducing ferroptosis in ectopic endometrial tissues of rats with endometriosis through the murine double minute 4 (MDM4)/tumor suppressor p53/glutathione peroxidase 4 (GPX4) signaling pathway. MethodsSeventy SPF-grade female SD rats were randomly divided into a blank group (n = 10), a sham-operated group (n = 10), and a modeling group (n = 50). The sham-operated group underwent laparotomy, while the modeling group was subjected to the autotransplantation method to establish an endometriosis model. After successful modeling, the animals were randomly assigned to the model group, progesterone group (0.25 mg·kg-1), and modified Shaofu Zhuyutang high-, medium-, and low-dose groups (30, 15, and 7.5 g·kg-1, respectively), with 10 rats per group. After four weeks of drug administration, the rats were euthanized for sample collection. The weight and volume of ectopic endometrial tissues were recorded for each group. Transmission electron microscopy (TEM) was employed to observe ultrastructural changes in endometrial tissues, while Prussian blue staining was used to assess iron ion deposition. Serum levels of interleukin-6 (IL-6), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α) were measured by enzyme-linked immunosorbent assay (ELISA). The relative levels of Fe2+, malondialdehyde (MDA), and glutathione (GSH) in endometrial tissues were determined by colorimetric assay. Immunofluorescence (IF) was used to detect the relative fluorescence intensities of GSH and GPX4 in endometrial tissues. The relative expression levels of phosphatidylinositol 3-kinase (PI3K), phosphorylated PI3K (p-PI3K), protein kinase B (Akt), phosphorylated Akt (p-Akt), MDM4, p53, and GPX4 proteins were detected by Western blot. Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) was used to assess the mRNA expression of PI3K, Akt, MDM4, p53, and GPX4. ResultsCompared with the blank and sham-operated groups, the model group exhibited reduced ferroptotic damage in ultrastructural observations, decreased ferroptotic aggregates and positive iron ion expression area on Prussian blue staining, elevated serum IL-6, IL-1β, and TNF-α levels, reduced Fe2+ and MDA content, increased GSH content in endometrial tissues, and enhanced GSH and GPX4 fluorescence intensities (P<0.01). The protein and mRNA expression levels of PI3K, p-PI3K, Akt, p-Akt, MDM4, and GPX4 were elevated, while those of p53 were decreased (P<0.01). Compared with the model group, in the progesterone group and the modified Shaofu Zhuyutang high- and medium-dose groups, ferroptotic damage in ultrastructural observations was exacerbated to varying degrees by TEM, and ferroptotic aggregates and positive iron ion expression areas were increased on Prussian blue staining. Serum IL-6, IL-1β, and TNF-α levels decreased, Fe2+ and MDA content increased, and GSH content decreased in endometrial tissues. GSH and GPX4 fluorescence intensities weakened, while the protein and mRNA expression levels of PI3K, p-PI3K, Akt, p-Akt, MDM4, and GPX4 decreased, and those of p53 increased (P<0.05, P<0.01). Conclusionmodified Shaofu Zhuyutang may exert therapeutic effects in endometriosis by inducing ferroptosis in ectopic endometrial tissues, alleviating inflammatory responses, and modulating key proteins in the MDM4/p53/GPX4 signaling pathway.


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