1.Exploring Biological Characteristics of Rat Model of Atrial Fibrillation with Phlegm-heat and Blood Stasis Pattern Based on Metabolomics
Ailin HOU ; Yuxuan LIU ; Wenxi YU ; Xing JI ; Chan WU ; Dazhuo SHI ; Ying ZHANG
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(5):245-255
ObjectiveTo establish an animal model of atrial fibrillation(AF) that accurately reflects the phlegm-heat and blood stasis(TRYZ) pathogenesis in traditional Chinese medicine. MethodsForty SPF-grade SD rats were randomly assigned using a random number table to the following groups:the control group, the TRYZ+AF group,the AF group and the TRYZ group, with ten rats in each group. The TRYZ+AF and TRYZ groups underwent a high-fat diet combined with intraperitoneal lipopolysaccharide(LPS) injection to simulate the pathological alterations of TRYZ syndrome. Groups TRYZ+AF and AF were induced with acetylcholine-calcium chloride(Ach-CaCl2) via caudal vein injection to induce AF. The control group received no intervention and was maintained under normal conditions. The modeling period lasted 3 weeks. Electrocardiography was used to assess AF episodes and duration, echocardiography evaluated left atrial dimensions and cardiac function, fully automated biochemical analyzer measured the levels of total cholesterol(TC), triglycerides(TG), high-density lipoprotein cholesterol(HDL-C) and low-density lipoprotein cholesterol(LDL-C), hemoreometer analyzed the whole blood viscosity, plasma viscosity, and whole blood reduced viscosity, a coagulation analyzer assessed prothrombin time(PT), activated partial thromboplastin time(APTT), thrombin time(TT), and fibrinogen(FIB), enzyme-linked immunosorbent assay(ELISA) was used to determine the levels of C-reactive protein(CRP), interleukin(IL)-1β, IL-6, IL-17, tumour necrosis factor(TNF)-α, matrix metalloproteinase-9(MMP-9), galectin-3(Gal-3), Collagen Ⅰ, and α-smooth muscle actin(α-SMA). Hematoxylin-eosin(HE) staining and Masson's trichrome staining were used to analyze pathological changes in atrial myocardium, Western blot was employed to detect MMP-9, Collagen Ⅰ and α-SMA protein expression in myocardial tissue, real-time quantitative polymerase chain reaction(Real-time PCR) evaluated fibrous factor gene expression levels. Changes in the TRYZ syndrome were assessed via body weight, tongue color[red(R), green(G), and blue(B)], and rectal temperature. Ultra-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry(UPLC-Q-TOF-MS) was employed to detect differential metabolites between the control group and the TRYZ+AF group. ResultsFollowing three weeks of sustained modeling, compared with the control group, rats in the TRYZ+AF and the TRYZ groups exhibited reduced body weight, dry faeces, elevated rectal temperature, dark red tongue, decreased RGB values on the tongue surface, and markedly elevated TC and LDL-C levels(P<0.05, P<0.01). The TRYZ+AF, TRYZ, and AF groups exhibited significantly decreased TT, APTT and PT, along with markedly elevated whole blood viscosity and FIB(P<0.05, P<0.01). Rats in the TRYZ+AF and AF groups exhibited AF rhythm, markedly decreased heart rate, prolonged RR intervals, enlarged left atrium, and significantly reduced ejection fraction and shortening fraction(P<0.05, P<0.01). Serum levels of CRP, IL-1β, IL-6, IL-17, TNF-α, MMP-9, Gal-3, Collagen Ⅰ, and α-SMA were elevated in rats from the TRYZ+AF, TRYZ, and AF groups compared to the control group, with the most pronounced increase observed in the TRYZ+AF group(P<0.05, P<0.01). Histopathology revealed that the collagen fiber deposition in the atrial of rats in the TRYZ+AF, TRYZ and AF groups was higher than that in the control group(P<0.05, P<0.01). Western blot and Real-time PCR results further demonstrated that the protein and mRNA expression levels of MMP-9, Collagen Ⅰ and α-SMA in the myocardial tissue of the TRYZ+AF group were higher than those in the other three groups(P<0.05, P<0.01). Metabolomic analysis revealed 173 differentially expressed metabolites in the TRYZ+AF group and the control group, primarily enriched in pathways such as glycerophospholipid metabolism and glycolysis/gluconeogenesis. ConclusionThis study successfully establishes a rat model of AF integrated with the TRYZ syndrome, demonstrating the pathological process where the interactions of phlegm, heat and stasis jointly trigger tremor, this provides a reliable experimental tool for in-depth research into the biological basis of this disease syndrome.
2.Mechanism of Dangui Shaoyaosan in Alleviating Inflammatory Responses in Diabetic Kidney Disease by Modulating Macrophage Polarization in Kidneys of db/db Mice
Luyu HOU ; Linlin ZHENG ; Wenjing SHI ; Zixuan WANG ; Shilong GUO ; Zhe LYU ; Dengzhou GUO
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(6):1-10
ObjectiveTo observe the effects of Danggui Shaoyaosan on macrophage polarization and renal inflammation in db/db mice with diabetic kidney disease (DKD), and to explore its renal protective effects and underlying mechanisms. MethodsEight db/m mice were assigned to the normal group, and forty db/db mice were randomly divided into a model group, low-, medium-, and high-dose Danggui Shaoyaosan groups (8.39, 16.77, 33.54 g·kg-1), and an irbesartan group (0.025 g·kg-1). All mice were administered treatment by gavage for 12 consecutive weeks. General conditions of the mice were observed during the intervention. At the end of the 12-week intervention, 24-h urine samples were collected using metabolic cages, after which the mice were anesthetized for sample collection. Blood was collected by enucleation and centrifuged to obtain serum for the determination of glycated serum protein (GSP), serum creatinine (SCr), blood urea nitrogen (BUN), total cholesterol (TC), and triglycerides (TG). The urinary albumin-to-creatinine ratio (UACR) was measured. Renal pathological changes were observed using hematoxylin-eosin (HE) staining, periodic acid-Schiff (PAS) staining, and Masson staining. Enzyme-linked immunosorbent assay (ELISA) was used to detect serum tumor necrosis factor-α (TNF-α), interleukin-10 (IL-10), and monocyte chemoattractant protein-1 (MCP-1) levels. Immunofluorescence (IF) was performed to detect F4/80 expression in renal tissue, and immunohistochemistry (IHC) was used to assess CD206 expression. Real-time quantitative polymerase chain reaction (Real-time PCR) was employed to measure the mRNA expression of TNF-α, IL-10, inducible nitric oxide synthase (iNOS), and arginase-1 (Arg-1). Western blot analysis was used to detect the protein expression of iNOS, Arg-1, CD86, and CD206 in renal tissue. ResultsCompared with the normal group, the model group showed increased levels of GSP, UACR, SCr, BUN, TC, and TG, elevated levels of the inflammatory factor TNF-α and the chemokine MCP-1, and decreased IL-10 levels (P<0.01). Pathological examination revealed glomerular hypertrophy, mesangial cell proliferation with marked mesangial expansion, inflammatory cell infiltration, vacuolar degeneration of renal tubular epithelial cells, prominent glycogen deposition, and increased collagen fiber deposition. In addition, relative F4/80 fluorescence intensity was enhanced, CD206 expression in the glomeruli and renal interstitium was reduced, and TNF-α and iNOS mRNA expression was increased. IL-10 and Arg-1 mRNA expression was decreased, iNOS and CD86 protein expression was increased, and Arg-1 and CD206 protein expression was decreased (P<0.05, P<0.01). Compared with the model group, the Danggui Shaoyaosan groups and the irbesartan group showed decreased levels of GSP, UACR, SCr, BUN, TC, and TG, reduced serum TNF-α and MCP-1 levels, and increased IL-10 levels. Renal pathological damage was improved to varying degrees. Relative F4/80 fluorescence intensity was reduced, CD206 expression in the glomeruli and renal interstitium was increased, and TNF-α and iNOS mRNA expression was decreased. IL-10 and Arg-1 mRNA expression was increased, iNOS and CD86 protein expression was reduced, and Arg-1 and CD206 protein expression was increased (P<0.05, P<0.01). ConclusionDanggui Shaoyaosan can improve renal function and alleviate renal pathological damage in db/db mice. Its mechanism may be related to inhibiting M1 pro-inflammatory macrophage polarization, promoting M2 anti-inflammatory macrophage polarization, reducing inflammatory responses, delaying the progression of renal fibrosis, improving renal pathological injury, and thereby exerting renal protective effects.
3.Mechanism of Danggui Shaoyaosan in Improving Inflammatory Response in Mice with Diabetic Kidney Disease Based on TLR4/p65/NLRP3 Signaling Pathway
Shilong GUO ; Ruijia LI ; Zixuan WANG ; Xinai WANG ; Luyu HOU ; Wenjing SHI ; Mengyuan TIAN ; Dengzhou GUO
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(6):19-27
ObjectiveTo investigate the effect of Danggui Shaoyaosan on the expression of Toll-like receptor 4/nuclear factor-kappa B p65/NOD-like receptor protein 3 (TLR4/NF-κB p65/NLRP3) signaling pathway in the renal tissues of db/db mice with spontaneous diabetes, and to explore the potential mechanism by which Danggui Shaoyaosan alleviates inflammation in diabetic kidney disease (DKD). MethodsThirty db/db mice were divided into five groups: A model group, Danggui Shaoyaosan low- (16.77 g·kg-1·d-1), medium- (33.54 g·kg-1·d-1), and high-dose (67.08 g·kg-1·d-1) intervention groups, as well as an irbesartan group (0.025 g·kg-1·d-1) by the random number table method, with 6 mice in each group. Additionally, 6 db/m mice were assigned to the normal group. After 8 weeks of intervention, the following parameters were determined by corresponding methods: body weight, fasting blood glucose (FBG), 24-hour urinary protein (24 h-UTP), and serum creatinine (SCr) levels, renal histopathological analysis by hematoxylin-eosin (HE) staining, Masson staining, and periodic acid-Schiff (PAS) staining, the protein and mRNA expression levels of TLR4, NF-κB p65, NLRP3, tumor necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-10 (IL-10), and interleukin-18 (IL-18) by Western blot and Real-time quantitative polymerase chain reaction (Real-time PCR), as well as TLR4, NF-κB p65, and NLRP3 protein expression in renal tissues by immunohistochemistry (IHC). ResultsCompared with the normal group, the model group exhibited increased body weight, FBG, 24 h-UTP, and SCr levels (P<0.05); disordered renal structure, thickened basement membrane, and interstitial inflammatory cell infiltration, elevated TLR4, NF-κB p65, NLRP3, TNF-α, IL-1β, IL-6, and IL-18 expression; as well as decreased IL-10 expression (P<0.05). Compared with the model group, these pathological changes and biochemical abnormalities were reversed in the medicine intervention groups to varying degrees (P<0.05). ConclusionDanggui Shaoyaosan may delay DKD progression by alleviating renal inflammatory response and reducing urinary protein excretion via modulating the TLR4/NF-κB p65/NLRP3 signaling pathway.
4.Factors influencing treatment outcomes and the construction of a prediction model following neoadjuvant endocrine therapy in patients with ER-positive breast cancer
ZHANG Hua1 ; XIE Shurui1 ; HOU Fengqian2 ; WANG Fei3 ; SHI Jianfei4
Chinese Journal of Cancer Biotherapy 2026;33(5):554-562
[摘 要] 目的:探究雌激素受体(ER)阳性乳腺癌患者新辅助内分泌治疗后病理完全缓解(pCR)的疗效影响因素,构建并验证预测模型。方法:回顾性选取2020年1月至2024年1月河北医科大学第四医院收治的715例接受新辅助内分泌治疗及手术治疗的ER阳性乳腺癌患者,根据术后病理结果分为pCR组(72例)和未达pCR组(643例)。按7∶3比例随机分为训练集和测试集,收集患者临床病理资料及实验室指标。通过单因素和多因素Logistic回归分析筛选未达pCR的独立危险因素,并构建预测模型。利用训练集和测试集的受试者工作特征(ROC)曲线、校准曲线及决策曲线分析(DCA)评估模型性能。结果:训练集与测试集患者临床资料比较无显著差异。训练集中,pCR组与未达pCR组在肿瘤增殖标志蛋白Ki-67指数、治疗前血清癌抗原15-3(CA15-3)水平、治疗前雌激素受体1(ESR1)基因突变状态、治疗前miR-1-3p表达水平、治疗前性别决定区Y框蛋白9(SOX9)表达水平、临床分期、淋巴结转移等方面存在差异(P < 0.05)。多因素Logistic回归分析显示,Ki-67指数、治疗前血清CA15-3水平、治疗前ESR1基因突变状态、治疗前miR-1-3p表达水平、临床分期为ER阳性乳腺癌患者新辅助内分泌治疗后未达pCR的独立影响因素(P < 0.05)。基于上述因素构建列线图预测模型,并通过ROC曲线、校准曲线及DCA验证模型具有较好的区分度、预测准确性和临床实用性。结论:基于Ki-67指数、血清CA15-3水平、ESR1基因突变状态等因素构建的列线图预测模型,对ER阳性乳腺癌患者新辅助内分泌治疗后未达到pCR风险具有良好预测效能,可为临床决策提供参考。
5.Expert consensus on precise intervention with repetitive transcranial magnetic stimulation for sleep disorders in the elderly
Yuan SHAO ; Jian WANG ; Wei LIANG ; Yingli ZHANG ; Gangqiang HOU ; Xia LI ; Yi XING ; Lu WANG ; Shi TANG ; Yongjun WANG
Sichuan Mental Health 2026;39(2):97-105
In recent years, repetitive transcranial magnetic stimulation (rTMS) has garnered significant attention as a therapeutic approach for sleep disorders in the elderly. However, the prevailing rTMS protocols are predominantly developed based on normative neurophysiological data derived from young adults and fail to incorporate individualized parameters tailored to the brain characteristics of the elderly. To address this gap, the consensus development group synthesized the latest evidence from 2010 to 2025 and established a standardized rTMS protocol specifically for elderly patients with sleep disorders. Adhering to the Appraisal of Guidelines for Research and Evaluation II (AGREE II) framework, systematically screened randomized controlled trials (RCTs) and systematic reviews regarding rTMS in the treatment of sleep disorders across various conditions. Meanwhile, the Grading of Recommendations Assessment, Development and Evaluation (GRADE) system was employed to rigorously grade the quality of evidence and the strength of recommendations. This consensus guideline delineates precise rTMS protocols for the management of sleep disorders in the elderly, highlights the adjustment of stimulation intensity according to scalp-cortex distance recommends either MRI‑guided neuronavigation or the Beam F3/F4 heuristic approach for accurate target localization, thereby providing precise rTMS intervention protocol for sleep disorders in the elderly, aiming to enhance clinical efficacy while ensuring treatment safety. [Funded by National Key Research and Development Program (number, 2023YFC3603200); General Program of Shenzhen Science and Technology Innovation Commission (number, JCYJ20240813112859008, JCYJ20240813112900002); Youth Program of Shenzhen Kangning Hospital (number, KN2023A004); www.guidelines-registry.cn number, PREPARE-2026CN530]
6.Review and reflection on the practice of pre-ethical review for drug clinical trials
Min HOU ; Xiaoyuan SHI ; Rongguo SUN ; Fang LIU ; Lei CHEN ; Zejin ZUO
Chinese Medical Ethics 2026;39(5):594-600
ObjectiveTo sort out the pre-ethical review status of drug clinical trials in a tertiary A hospital, summarize practical experience, and provide references for pre-ethical review. MethodA retrospective analysis was conducted on the ethical review of pre-ethical projects in a tertiary A hospital from 2018 to 2023. ResultsFrom 2018 to 2023, a total of 285 pre-ethical projects were reviewed in this tertiary hospital, including 79 projects where it served as the leading unit. Among these, 279 clinical trials ultimately received approval of the ethical review committee, while 6 projects were not approved due to sponsors’ refusal to modify study protocols. In terms of trial phases, phase III clinical trials constituted the largest proportion, and the oncology center was the department with the highest number of projects. In 2023, the average time for pre-ethical review projects from submission to approval was 43 calendar days, 3 days longer than for other project types. In 2023, this hospital reviewed 75 pre-ethical review projects, including 58 where it served as a participating unit. Among these, 42 projects received approval later than the leading unit, while 9 projects were approved earlier than the leading unit’s ethical approval date. Among the pre-ethical review projects applied in 2023, 70 projects obtained drug clinical trial notifications from the National Medical Products Administration, while 5 projects had unknown notification status due to the lack of ethical approval or discontinuation. Of the projects receiving approval notifications, 16 were annotated with matters requiring enhanced attention during clinical trials, and 7 necessitated protocol improvements. ConclusionThis tertiary A hospital has implemented multiple measures to optimize the management of pre-ethical review. This ethical review model does not compromise the quality of ethical review and contributes to accelerating the initiation of clinical trials. Notably, it is crucial to construct a patient-centered drug development system. Clinical trials guided by this concept align with ethical values, laying the foundation for the smooth conduct of clinical trials, assisting in the drug development process, and safeguarding patients’ medication needs.
7.Feasibility of 4-minute respiratory-gated total-body PET/CT in patients with malignancies
Xiaoguang HOU ; Shuguang CHEN ; Yimeng SHI ; Haojun YU ; Pengcheng HU ; Guobing LIU
Chinese Journal of Clinical Medicine 2026;33(3):452-460
Objective To explore the feasibility and image quality of respiratory-gated total-body 18F-FDG PET/CT imaging with a 4-minute acquisition in patients with malignant tumors. Methods Sixty-two patients with pathologically confirmed malignant tumors (118 lesions measured) who underwent total-body 18F-FDG PET/CT examination were consecutively enrolled at the Department of Nuclear Medicine, Zhongshan Hospital, Fudan University, from September 23, 2021 to November 23, 2021, and their clinical and imaging data were retrospectively collected. PET data was acquired over 12 minutes with simultaneous respiratory waveform recorded. The data were reconstructed into four image sets: gated-12 group and gated-4 group (using all counts and the first 4-minute counts, respectively) and ungated-12 group and ungated-6 group (using all counts and the first 6-minute counts, respectively) reconstructed with respiratory gating that retained counts corresponding to the half respiratory window near end-expiration. Image quality assessment and quantitative analysis of lesions were performed and compared among the four image sets, and subgroup analyses were performed according to lesion location, size, and metabolic activity. Results The effective counts of 4 sets of PET images were highest in ungated-12 group, followed by ungated-6 group and gated-12 group, and lowest in gated-4 group (P<0.001). Regardless of the metabolic level and size of the lesions, compared with the ungated-12 group and the ungated-6 group, the lesions in the gated-12 group showed higher maximum, average, and peak standardized uptake values (SUVmax, SUVmean, and SUVpeak), tumor liver uptake ratio, tumor blood pool uptake ratio, and smaller metabolic volume (P<0.05); There was no statistically significant difference in the above indicators between the gate-12 group and the gate-4 group. The background noise of gate-4 group images is slightly higher than that of gate-12 group images (Liver SUVSD: [0.17+0.05] vs [0.14+0.04]); Blood pool SUVSD: (0.08 ± 0.03) vs (0.06 ± 0.03). There was a statistically significant difference in subjective scores of PET image quality among the four groups (P<0.001). The subjective scores of image quality in the gated-12 group (4.79 ± 0.41) were not significantly different from those in the non gated-12 group (4.97 ± 0.17) and non gated-6 group (4.79 ± 0.45), but were higher than those in the gated-4 group ([4.17 ± 0.45], P<0.001), And the scores have good consistency (Kappa>0.75). Conclusion In total-body PET/CT imaging for malignant tumors, 4-minute respiratory-gated acquisition can achieve comparable quantitative performance to longer respiratory gating imaging while significantly shortening acquisition time. This approach demonstrates promising potential for clinical application.
8.Comparison of efficacy and safety of mizoribine and mycophenolate mofetil in kidney transplant recipients: a single-center retrospective study
Xinji YANG ; Weilong SHI ; Zaiwei SONG ; Zhifei XIE ; Herong ZHU ; Wenbin ZHANG ; Hongxian ZHANG ; Lei LIU ; Lei ZHAO ; Lu WANG ; Zhidan WANG ; Shudong ZHANG ; Qiming ZHANG ; Xiaofei HOU
Organ Transplantation 2026;17(4):625-634
Objective To compare the clinical efficacy and safety differences between the triple immunosuppressive regimen of mizoribine (MZR) or mycophenolate mofetil (MMF) combined with tacrolimus (Tac) and glucocorticoids of kidney transplant recipients in one year after transplantation. Methods A single-center retrospective cohort study design was adopted. The clinical data of 156 patients who underwent the first allogeneic kidney transplantation at the Third Hospital of Peking University from January 2022 to December 2024 were included. The patients were divided into the MZR group (78 cases) and the MMF group (78 cases) based on the initial immunosuppressive regimen after transplantation. The baseline data, medication use in one year after transplantation, blood routine indicators, adverse events and clinical outcomes were compared between the two groups. For the repeated-measurement longitudinal data, repeated-measurement analysis of variance was used to evaluate the main effects of the groups, time and their interaction effects. Results At the primary efficacy endpoint, there were no statistically significant differences in the one-year survival rate, graft survival rate and incidence of acute rejection between the two groups (all P > 0.05). In terms of medication patterns and laboratory test indicators, there were significant time main effects and "time × group" interaction effects for Tac dosage (all P < 0.05). The Tac dosage in the MZR group was lower than that in the MMF group from 2 to 3 months after transplantation, and it was higher in the MZR group than in the MMF group from 7 to 12 months and one year after transplantation. White blood cell count in the MZR group was higher at one month after transplantation, and the platelet count was higher one year after transplantation (all P < 0.05). In terms of infection, the incidence of novel coronavirus and Pneumocystis jirovecii pneumonia infections in the MZR group was lower (all P < 0.05). In terms of metabolic indicators, the post-transplantation uric acid level was better in the MZR group than in the MMF group, but the total cholesterol level was higher one year after transplantation (all P < 0.05). In terms of liver function, there was a brief and mild increase in transaminase in the early stage after transplantation in the MZR group. Conclusions During the one-year follow-up after transplantation, both the MZR and MMF regimens demonstrate comparable immunosuppressive efficacy and overall safety in Chinese kidney transplant recipients. The MZR regimen shows clear advantages in reducing specific infection risks, alleviating bone marrow suppression and improving uric acid metabolism, but its potential impact on blood lipids needs to be monitored. Therefore, the MZR regimen may be an effective and safe alternative immunosuppressive option for kidney transplant recipients, especially those with high infection risk, poor hematological tolerance, or comorbid hyperuricemia.
9.The mechanism of magnoflorine in inhibiting colon cancer based on network pharmacology and in vitro experiment
Lulu TAN ; Lina ZHU ; Shujin ZHANG ; Yuxuan WANG ; Huimei LI ; Yuke WANG ; Jiayi HOU ; Qilong FENG ; Jianyun SHI
Acta Universitatis Medicinalis Anhui 2026;61(6):1021-1031
ObjectiveTo explore the function and related molecular mechanisms of magnoflorine against colon cancer via network pharmacology, molecular docking, and in vitro cell experiments. MethodsIn this study, the canonical SMILES of magnoflorine was obtained from the PubChem database, and the potential targets of magnoflorine were predicted by the Swiss Target Prediction database, while the disease targets of colon cancer were obtained from the DisGeNET, GeneCards and OMIM databases. The intersecting targets between magnoflorine's predicted targets and colon cancer disease targets were taken, and a protein-protein interaction (PPI) network was constructed and analyzed. The DAVID online database was employed to conduct Gene Ontology (GO) and KEGG pathway enrichment analyses on core targets. The top five key targets screened were docked with magnoflorine using AutoDock software. Finally, cellular experiments including CCK-8 assays, EdU experiments, cell scratch assays, and Transwell assays were conducted to validate the results from network pharmacology and molecular docking. Results44 key targets of magnoflorine in resisting colon cancer were acquired. The molecular docking results showed that magnoflorine had a strong binding activity with the core target signal transducer and activator of transcription 3 (STAT3) in the top five of the PPI network. Cellular experiments confirmed that magnoflorine could inhibit the proliferation and migration of colon cancer cells by suppressing the JAK/STAT3 signaling pathway. ConclusionMagnoflorine may inhibit the proliferation and migration of colon cancer cells by regulating the JAK/STAT3 signaling pathway.
10.Mechanism of Herbal Cake-separated Moxibustion in Improving Neuroimmune Inflammation in Rats with Chronic Fatigue Syndrome by Interfering TLR4/MyD88/NF-κB Pathway
Chuntao ZHAI ; Yawei HOU ; Linjuan SHI ; Yixiao WANG ; Wei LI ; Yuefeng TIAN
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(4):140-149
ObjectiveTo explore the mechanism of herbal cake-separated moxibustion using the classical formula Xiaoyaosan in alleviating neuroimmune inflammatory responses in chronic fatigue syndrome (CFS) rats, based on the regulation of the toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88)/nuclear transcription factor-κB (NF-κB) signaling pathway. MethodsFifty SPF-grade SD rats aged 6-8 weeks were randomly divided into five groups: Normal group, model group, sham herbal cake moxibustion group, Chinese medicine intragastric administration group, and herbal cake-separated moxibustion group, with 10 rats in each group. Except for the normal group, all other groups underwent a 21-day modeling process, followed by behavioral testing. The herbal cake-separated moxibustion and sham herbal cake moxibustion groups received interventions at the Shenque (CV8), Guanyuan (CV4), Zusanli (ST36), and Qimen (LR14) acupoints. The Chinese medicine intragastric administration group was treated with a Xiaoyaosan suspension via gavage. Behavioral tests were conducted after 10 days of continuous intervention. Serum levels of interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α), as well as hippocampal levels of IL-1β, IL-6, TNF-α, and NF-κB, were detected by enzyme-linked immunosorbent assay (ELISA). Morphological changes in the hippocampus were observed using hematoxylin-eosin (HE) staining. Real-time quantitative polymerase chain reaction (Real-time PCR) was used to detect mRNA expression levels of TLR4, MyD88, and NF-κB in the hippocampus. Western blot analysis was performed to detect the relative expression levels of TLR4, MyD88, NF-κB, and p65 proteins in the hippocampus. ResultsCompared with the normal group, the model group showed a significant decrease in upright times during the open field test (P<0.01), as well as significant reductions in total movement distance, resting time, and center region duration (P<0.01). In the tail suspension test, immobility time increased (P<0.01), and struggle times decreased (P<0.01). Serum and hippocampal levels of IL-1β, IL-6, and TNF-α, as well as hippocampal NF-κB levels and TLR4, MyD88, and NF-κB mRNA expression, were significantly elevated (P<0.01). After treatment, compared with the model group, the total movement distance and upright times in the open field test were significantly increased in all treatment groups (P<0.01), while resting time and center region duration were notably prolonged (P<0.05, P<0.01). Immobility time in the tail suspension test was significantly shortened (P<0.01), and struggle times significantly increased (P<0.05). Serum and hippocampal levels of IL-1β, IL-6, TNF-α, hippocampal NF-κB levels, and TLR4 and NF-κB mRNA expression were significantly reduced (P<0.05, P<0.01). Compared with the sham herbal cake moxibustion group, the herbal cake-separated moxibustion group showed a significant extension in center region duration during the open field test (P<0.05) and a significant increase in upright times (P<0.01). In the tail suspension test, immobility time was reduced (P<0.01), and struggle times increased (P<0.01). Serum TNF-α levels in the Chinese medicine intragastric administration group were significantly reduced (P<0.01), while serum IL-6 levels, as well as hippocampal levels of IL-1β, TNF-α, NF-κB, and TLR4, MyD88, and NF-κB mRNA expression, were significantly decreased in both the Chinese medicine intragastric administration group and the herbal cake-separated moxibustion group (P<0.05, P<0.01). Compared with the Chinese medicine intragastric administration group, the herbal cake-separated moxibustion group exhibited significantly increased upright times in the open field test (P<0.01). In the tail suspension test, immobility time was reduced (P<0.01), and struggle times increased (P<0.01). Serum IL-1β, hippocampal TNF-α levels, and TLR4, MyD88, and NF-κB mRNA expression were significantly decreased (P<0.05, P<0.01). ConclusionHerbal cake-separated moxibustion effectively improves fatigue and memory function in CFS rats, regulates neuroimmune inflammatory responses, and its mechanism may be related to the modulation of the TLR4/MyD88/NF-κB signaling pathway.

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