1.Mechanism of Shenfu Xiongze Prescription in Regulating Autophagy Level to Intervene in Myocardial Remodeling in Rats via AMPK/mTOR Signaling Pathway
Xueqing WANG ; Wei ZHONG ; Liangliang PAN ; Caihong LI ; Man HAN ; Xiaowei YANG ; Yuanwang YU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(2):136-144
ObjectiveTo explore the mechanism by which the Shenfu Xiongze prescription regulates autophagy in rats with myocardial remodeling through the adenosine monophosphate-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway. MethodsA rat model of myocardial remodeling induced by isoprenaline (ISO) was established. Rats were divided into the blank group,the model group,the low-,medium-, and high-dose groups of Shenfu Xiongze prescription,and the captopril group, 6 rats in each group. Except for the blank group,the rat model of myocardial remodeling was established in the other groups by intraperitoneal injection of 2.5 mg·kg-1 ISO for 3 consecutive weeks. At the same time of modeling, the low-,medium-, and high-dose groups of Shenfu Xiongze prescription were administered the corresponding doses of Shenfu Xiongze prescription solution (8.4,16.8,and 33.6 g·kg-1),and the captopril group was administered captopril solution (25 mg·kg-1). As for the blank group and the model group, the same volume of normal saline was given. The treatment was continued for 3 weeks. Echocardiography was used to observe the cardiac structure and function,and the heart weight index was detected. Masson staining and hematoxylin-eosin (HE) staining were used to observe the pathological morphology changes of myocardial tissue. The levels of interleukin-6 (IL-6) and B-type natriuretic peptide (BNP) in serum were detected by enzyme-linked immunosorbent assay (ELISA). The expression of type Ⅰ collagen (Collagen Ⅰ),type Ⅲ collagen (Collagen Ⅲ),and microtubule-associated protein 1 light chain 3 (LC3) proteins in myocardial tissue was determined by immunohistochemistry. Autophagy was observed by transmission electron microscopy. The mRNA expression of Collagen Ⅰ,Collagen Ⅲ,α-smooth muscle actin (α-SMA),LC3,yeast Atg6 homolog protein (Beclin-1),AMPK,and mTOR in myocardial tissue was detected by quantitative real-time polymerase chain reaction (real-time PCR). The protein expression of Collagen Ⅰ,α-SMA,transforming growth factor-β1 (TGF-β1),LC3,Beclin-1,p62, phosphorylation(p)-AMPK,p-mTOR,AMPK,and mTOR was detected by Western blot. ResultsCompared with the normal group,rats in the model group exhibited significantly decreased values of ejection fraction (EF) and left ventricular fractional shortening (FS) (P<0.01), significantly increased values of left ventricular end-diastolic diameter (LVIDd) and left ventricular end-systolic diameter (LVIDs) (P<0.01). Additionally, the model group also showed increased degrees of inflammatory infiltration and fibrosis of myocardial tissue, significantly elevated levels of serum IL-6 and BNP (P<0.01), significantly increased mRNA and protein levels of Collagen Ⅰ,Collagen Ⅲ,α-SMA,and mTOR (P<0.01),and markedly decreased mRNA and protein levels of LC3,Beclin-1,and AMPK (P<0.05,P<0.01). Compared with the model group, the low-,medium-, and high-dose groups of Shenfu Xiongze prescription presented significantly elevated EF and FS values (P<0.01) and lowered LVIDd and LVIDs (P<0.05). In these groups, the inflammation and fibrosis were alleviated significantly. They also exhibited decreased serum levels of IL-6 and BNP (P<0.01), significantly reduced protein expression of Collagen Ⅰ, α-SMA, TGF-β1, p62, and p-mTOR (P<0.01), significantly decreased mRNA expression of Collagen Ⅰ, Collagen Ⅲ, α-SMA, and mTOR (P<0.01), and significantly increased mRNA and protein levels of LC3, Beclin-1, and AMPK (P<0.05,P<0.01). ConclusionThe Shenfu Xiongze prescription can improve the myocardial remodeling induced by ISO in rats by regulating the autophagy level,enhance cardiac function,and reduce inflammatory and fibrotic levels. This effect may be achieved through the AMPK/mTOR signaling pathway.
2.Sanren Runchang Formula Regulates Brain-gut Axis to Treat IBS-C: A Randomized Controlled Trial
Teng LI ; Xinrong FAN ; He YAN ; Zhuozhi GONG ; Mengxi YAO ; Na YANG ; Yuhan WANG ; Huikai HU ; Wei WEI ; Tao LIU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(2):154-161
ObjectiveTo observe the clinical efficacy of Sanren Runchang formula in treating constipation-predominant irritable bowel syndrome (IBS-C) by regulating the brain-gut axis and the effects of the formula on serum levels of 5-hydroxytryptamine (5-HT), vasoactive intestinal peptide (VIP), and substance P (SP). MethodsA randomized controlled design was adopted, and 72 IBS-C patients meeting Rome Ⅳ criteria were randomized into observation and control groups (36 cases).The observation group received Sanren Runchang formula granules twice daily, and the control group received lactulose oral solution daily for 4 weeks. IBS Symptom Severity Scale (IBS-SSS), IBS Quality of Life Scale (IBS-QOL), and Bristol Stool Form Scale (BSFS) were used to assess clinical symptoms, and bowel movement frequency was recorded. The Self-Rating Anxiety Scale (SAS) and Self-Rating Depression Scale (SDS) were employed to evaluate psychological status. ELISA was employed to measure the serum levels of 5-HT, VIP, and SP. ResultsThe total response rate in the observation group was 91.67% (33/36), which was higher than that (77.78%, 28/36) in the control group (χ2=4.50, P<0.05). After treatment, both groups showed increased defecation frequency and BSFS scores, decreased IBS-SSS total score, abdominal pain and bloating scores, IBS-QOL health anxiety, anxiety, food avoidance, and behavioral disorders scores, SAS and SDS scores, serum 5-HT and VIP levels, and increased SP levels (P<0.05, P<0.01). Moreover, the observation group showed more significant changes in the indicators above than the control group (P<0.05, P<0.01). The SP level showed no significant difference between the two groups. During the 4-week follow-up, the recurrence rate was 5.88% in the observation group and 31.25% in the control group. No adverse events occurred in observation group, and 2 cases of mild diarrhea occurred in the control group. ConclusionSanren Runchang formula demonstrated definitive efficacy in alleviating gastrointestinal symptoms and improving the psychological status and quality of life in IBS-C patients, with a low recurrence rate. The formula can regulate serum levels of neurotransmitters such as 5-HT and VIP, suggesting its potential regulatory effect on the brain-gut axis through modulating neurotransmitters and neuropeptides. However, its complete mechanism of action requires further investigation through detection of additional brain-gut axis-related biomarkers.
3.Quality Evaluation of Naomaili Granules Based on Multi-component Content Determination and Fingerprint and Screening of Its Anti-neuroinflammatory Substance Basis
Ya WANG ; Yanan KANG ; Bo LIU ; Zimo WANG ; Xuan ZHANG ; Wei LAN ; Wen ZHANG ; Lu YANG ; Yi SUN
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(2):170-178
ObjectiveTo establish an ultra-performance liquid fingerprint and multi-components determination method for Naomaili granules. To evaluate the quality of different batches by chemometrics, and the anti-neuroinflammatory effects of water extract and main components of Naomaili granules were tested in vitro. MethodsThe similarity and common peaks of 27 batches of Naomaili granules were evaluated by using Ultra performance liquid chromatography (UPLC) fingerprint detection. Ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) technology was used to determine the content of the index components in Naomaili granules and to evaluate the quality of different batches of Naomaili granules by chemometrics. LPS-induced BV-2 cell inflammation model was used to investigate the anti-neuroinflammatory effects of the water extract and main components of Naomaili granules. ResultsThe similarity of fingerprints of 27 batches of samples was > 0.90. A total of 32 common peaks were calibrated, and 23 of them were identified and assigned. In 27 batches of Naomaili granules, the mass fractions of 14 components that were stachydrine hydrochloride, leonurine hydrochloride, calycosin-7-O-glucoside, calycosin,tanshinoneⅠ, cryptotanshinone, tanshinoneⅡA, ginsenoside Rb1, notoginsenoside R1, ginsenoside Rg1, paeoniflorin, albiflorin, lactiflorin, and salvianolic acid B were found to be 2.902-3.498, 0.233-0.343, 0.111-0.301, 0.07-0.152, 0.136-0.228, 0.195-0.390, 0.324-0.482, 1.056-1.435, 0.271-0.397, 1.318-1.649, 3.038-4.059, 2.263-3.455, 0.152-0.232, 2.931-3.991 mg∙g-1, respectively. Multivariate statistical analysis showed that paeoniflorin, ginsenoside Rg1, ginsenoside Rb1 and staphylline hydrochloride were quality difference markers to control the stability of the preparation. The results of bioactive experiment showed that the water extract of Naomaili granules and the eight main components with high content in the prescription had a dose-dependent inhibitory effect on the release of NO in the cell supernatant. Among them, salvianolic acid B and ginsenoside Rb1 had strong anti-inflammatory activity, with IC50 values of (36.11±0.15) mg∙L-1 and (27.24±0.54) mg∙L-1, respectively. ConclusionThe quality evaluation method of Naomaili granules established in this study was accurate and reproducible. Four quality difference markers were screened out, and eight key pharmacodynamic substances of Naomaili granules against neuroinflammation were screened out by in vitro cell experiments.
4.Joint Relation Extraction of Famous Medical Cases with CasRel Model Combining Entity Mapping and Data Augmentation
Yuxin LI ; Xinghua XIANG ; Hang YANG ; Dasheng LIU ; Jiaheng WANG ; Zhiwei ZHAO ; Jiaxu HAN ; Mengjie WU ; Qianzi CHE ; Wei YANG
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(2):218-225
ObjectiveTo address the challenges of unstructured classical Chinese expressions, nested entity relationships, and limited annotated data in famous traditional Chinese medicine(TCM) case records, this study proposes a joint relation extraction framework that integrates data augmentation and entity mapping, aiming to support the construction of TCM diagnostic knowledge graphs and clinical pattern mining. MethodsWe developed an annotation structure for entities and their relationships in TCM case texts and applied a data augmentation strategy by incorporating multiple ancient texts to expand the relation extraction dataset. A cascade binary tagging framework for relation triple extraction(CasRel) model for TCM semantics was designed, integrating a pre-trained bidirectional encoder representations from transformers(BERT) layer for classical TCM texts to enhance semantic representation, and using a head entity-relation-tail entity mapping mechanism to address entity nesting and relation overlapping issues. ResultsExperimental results showed that the CasRel model, combining data augmentation and entity mapping, outperformed the pipeline-based Bert-Radical-Lexicon(BRL)-bidirectional long short-term memory(BiLSTM)-Attention model. The overall precision, recall, and F1-score across 12 relation types reached 65.73%, 64.03%, and 64.87%, which represent improvements of 14.26%, 7.98%, and 11.21% compared to the BRL-BiLSTM-Attention model, respectively. Notably, the F1-score for tongue syndrome relations increased by 22.68%(69.32%), and the prescription-syndrome relations performed the best with the F1-score of 70.10%. ConclusionThe proposed framework significantly improves the semantic representation and complex dependencies in TCM texts, offering a reusable technical framework for structured mining of TCM case records. The constructed knowledge graph can support clinical syndrome differentiation, prescription optimization, and drug compatibility, providing a methodological reference for TCM artificial intelligence research.
5.Comparison of preoperative ocular biometry between Pentacam AXL and IOL Master 700 in cataract patients
Jinfen WEI ; Simin TAN ; Lin DING ; Qiuli ZHANG
International Eye Science 2026;26(1):148-151
AIM: To compare the preoperative ocular biometry between Pentacam AXL and IOL Master 700 in cataract patients.METHODS:Prospective study. A total of 150 patients(150 eyes)with cataracts who were treated in our hospital from May to December 2024 were selected. The IOL Master 700 and Pentacam AXL were preoperatively used to measure axial length(AL), corneal curvature(K1, K2 and Km), anterior chamber depth(ACD), and white-to-white(WTW). The difference and consistency of the results of the two instruments were compared.RESULTS: There was no significant difference between the two instruments in the AL, K1, K2, Km, ACD, and WTW(all P>0.05). The Spearman correlation analysis showed that the two instruments positively correlated with the AL, K1, K2, Km, ACD and WTW of the operated eye(all P<0.001). The Bland-Altman analysis showed that for the Pentacam AXL and IOL Master 700, there were 5/150(3.3%), 7/150(4.7%), 4/150(2.7%), 5/150(3.3%), and 0 points outside the 95%LoA for the AL, K1, K2, Km, ACD, and WTW of the examined eyes, respectively, with all of these values less than 5%, indicating good consistency.CONCLUSION:The AL, K1, K2, Km, ACD and WTW of Pentacam AXL and IOL Master 700 in cataract patients before cataract phacoemulsification combined with IOL implantation show no significant differences, and have good correlation and consistency. The two instruments can be used interchangeably.
6.Neuroprotective Mechanism of Yifei Xuanfei Jiangzhuo Prescription on VaD Rats Based on NF-κB/NLRP3 Signaling Pathway
Bingmao YUAN ; Wei CHEN ; Xiu LAN ; Lingfei JIANG ; Lin WU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(3):88-96
ObjectiveTo investigate the molecular mechanism by which Yifei Xuanfei Jiangzhuo prescription regulates the nuclear factor-κB (NF-κB)/NOD-like receptor protein 3 (NLRP3) signaling pathway to improve neuronal function in vascular dementia (VaD) rats. MethodsA VaD model was established by intermittently clamping the bilateral common carotid arteries (CCA) combined with bilateral vascular occlusion (2-VO). Eighty-four SD rats were randomly divided into a blank group, sham group, model group, piracetam group (0.2 g·kg-1), and low-, medium-, and high-dose Yifei Xuanfei Jiangzhuo prescription groups (6.09, 12.18, and 24.36 g·kg-1). Drug administration began on day 7 after surgery, once daily for 28 consecutive days. Behavioral experiments were used to evaluate learning and spatial memory. Hematoxylin-eosin (HE) staining was applied to observe pathological morphological changes in the CA1 region of the hippocampus. Transmission electron microscopy was used to examine the ultrastructure of hippocampal neurons. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) was used to detect neuronal apoptosis in the CA1 region. Immunohistochemistry was performed to determine the positive expression rate of neuronal nuclear antigen (NeuN). Immunofluorescence single staining was used to assess nuclear expression of NF-κB p65 in brain tissue. Western blot was used to detect the protein expression levels of inhibitor of κB kinase (IKK), NF-κB p65, NLRP3, Caspase-1, apoptosis-associated speck-like protein (ASC), and interleukin-1β (IL-1β). ResultsCompared with the blank group, the model group showed a significant reduction in platform-crossing frequency (P0.01), aggravated hippocampal injury, a significant increase in neuronal apoptosis (P0.05), decreased NeuN positivity in the CA1 region (P0.05), increased nuclear expression of NF-κB p65 (P0.05), and significantly elevated expression of p-IKK, p-NF-κB p65, NLRP3, cleaved Caspase-1, ASC, and cleaved IL-1β (P0.05). Compared with the model group, all drug-treated groups improved learning and spatial memory in VaD rats, alleviated hippocampal pathological injury and neuronal apoptosis, and protected neuronal ultrastructure. Yifei Xuanfei Jiangzhuo prescription at doses of 12.18 and 24.36 g·kg-1 reduced hippocampal expression levels of p-IKK, p-NF-κB p65, NLRP3, Caspase-1, ASC, and cleaved IL-1β in VaD rats (P0.05), showing dose-dependent inhibition of the NF-κB/NLRP3 signaling pathway. ConclusionYifei Xuanfei Jiangzhuo prescription may exert neuroprotective effects by regulating the NF-κB/NLRP3 signaling pathway, thereby reducing neuroinflammation and inhibiting hippocampal neuronal apoptosis.
7.Exploring Mechanism of Yiqi Huoxue Jiedu Formula in Alleviating Immune Cell Exhaustion in Sepsis Based on Transcriptomics and Metabolomics
Rui CHEN ; Qiusha PAN ; Kaiqiang ZHONG ; Shuqi MA ; Wei HUANG ; Jiahua LAI ; Ruifeng ZENG ; Xiaotu XI ; Jun LI
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(3):109-118
ObjectiveTo observe the effects of Yiqi Huoxue Jiedu formula(YHJF) on immune cell exhaustion in the spleen of septic mice and to explore and validate its potential intervention targets. MethodsMice were randomly divided into the sham-operated, model, low-dose YHJF(4.1 g·kg-1), and high-dose YHJF(8.2 g·kg-1) groups. Except for the sham-operated group, a cecal ligation and puncture(CLP) procedure was performed to establish a mouse sepsis model. The treatment groups received oral administration of the corresponding doses, while the sham-operated and model groups received an equal volume of physiological saline. After the intervention, the 7-day survival rate of each group was recorded, and spleen samples were collected 72 h post-intervention, and the spleen index was calculated. Terminal deoxynucleotidyl transferase deoxyuridine triphosphate(dUTP) nick end labeling(TUNEL) staining was used to detect apoptosis in spleen cells. Enzyme-linked immunosorbent assay(ELISA) was performed to measure the levels of interleukin(IL)-4 and IL-10 in the serum. Transcriptomics and metabolomics were used to screen for differentially expressed genes(DEGs) and differential metabolites in the spleen, followed by bioinformatics analysis to identify key targets. Real-time quantitative polymerase chain reaction(Real-time PCR), flow cytometry, and multiplex immunofluorescence were used to verify the expressions of key genes and proteins. ResultsThe high-dose YHJF group significantly improved the 7-day survival rate of septic mice(P0.05). Compared with the sham-operated group, the model group showed a significant increase in apoptosis of spleen cells and a decrease in the spleen index at 72 h post-modeling, with markedly elevated peripheral serum IL-4 and IL-10 levels(P0.01). Compared with the model group, the high-dose YHJF group showed a reduction in apoptosis of spleen cells, an increase in the spleen index, and a significant decrease in peripheral serum IL-4 and IL-10 levels(P0.05). Spleen transcriptomics identified 255 DEGs between groups, potentially serving as intervention targets for YHJF. Gene Ontology(GO) enrichment analysis revealed that DEGs were mainly involved in biological processes such as natural killer(NK) cell-mediated positive immune regulation, cell killing, cytokine production, positive regulation of innate immune cells, and interferon production. Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analysis showed that DEGs were mainly involved in cytokine-cytokine receptor interactions, viral protein interactions with cytokines and cytokine receptors, chemokine signaling pathway, and nuclear transcription factor-κB(NF-κB) signaling pathway. Protein-protein interaction(PPI) network analysis identified CD160, granzyme B(GZMB), and chemokine ligand 4(CCL4) as key targets for YHJF in treating sepsis. Metabolomics identified 46 differential metabolites that were significantly reversed by YHJF intervention, and combined transcriptomics and metabolomics analysis identified 17 differential metabolites closely related to CD160. Pathway enrichment revealed that these metabolites were mainly involved in glycerophospholipid metabolism, arachidonic acid metabolism, glycosylphosphatidylinositol(GPI) anchor biosynthesis, linoleic acid metabolism, and α-linolenic acid metabolism pathways. Verification results showed that, compared with the sham-operated group, the model group exhibited significantly elevated CD160 mRNA expression level in the spleen, along with markedly decreased CCL4 and GZMB mRNA expression, and had a significant increase in CD160 expression on the surface of natural killer T(NKT) cells in the spleen(P0.01). Compared with the model group, the high-dose YHJF group had a significant decrease in CD160 mRNA expression in the spleen, a significant increase in CCL4 and GZMB mRNA expressions. Further flow cytometry and immunofluorescence revealed that compared with the sham-operated group, CD160 expression on the surface of splenic NKT cells in the model group was significantly increased(P0.01), while high-dose YHJF intervention significantly reduced CD160 expression(P0.01). ConclusionYHJF may alleviate NKT cell exhaustion in sepsis by downregulating the expression of the negative co-stimulatory molecule CD160, and this regulatory effect is closely related to fatty acid metabolism pathways. This study provides new insights and targets for further exploration of strengthening vital Qi and detoxifying strategy to improve immune cell exhaustion in acute deficiency syndrome of sepsis.
8.Xuefu Zhuyutang in Malignant Tumor Disease: A Review
Jiaqi JI ; Xiaoqing HU ; Yihan ZHAO ; Xuhang SUN ; Dandan WEI ; Junwen PEI ; Shiqing JIANG
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(3):321-330
Cancer has become a significant global public health issue, severely impacting public health and societal development. Despite advances in tumor treatment methods in recent years and a gradual decline in cancer mortality rates, drug-related adverse reactions and drug resistance remain substantial challenges. Traditional Chinese medicine (TCM) has demonstrated significant clinical efficacy in cancer treatment and small side effects, making it widely applied in the field of oncology. Xuefu Zhuyutang, derived from Yilin Gaicuo, is known for its abilities to invigorate blood circulation, dispel blood stasis, promote Qi flow, and alleviate pain. It was specifically formulated by the esteemed WANG Qingren of the Qing dynasty for the "blood stasis syndrome in the blood mansion" and is commonly used to treat Qi stagnation and blood stasis syndrome. Clinical studies have shown that Xuefu Zhuyutang, when combined with conventional Western medications, produces significant effects in the treatment of malignant tumors such as liver cancer, lung cancer, and cervical cancer. It substantially reduces the incidence of adverse reactions following Western treatments, including radiation esophagitis, radiation encephalopathy, radiation-induced oral mucositis, and edema. Additionally, it alleviates cancer-related pain and fever, blood hypercoagulability, and associated complications such as depression and anxiety, and also mitigates chemotherapy-induced side effects like hand-foot syndrome. Basic research has demonstrated its potential anti-tumor mechanisms, including the inhibition of Wnt/β-catenin signaling pathway activation, suppression of mitogen-activated protein kinase (MAPK) pathway activation, and anti-tumor angiogenesis. Pharmacological studies have revealed that its active components inhibit tumor cell proliferation and migration, induce tumor cell apoptosis, suppress tumor angiogenesis, enhance the cytotoxicity of natural killer cells against tumors, improve the tumor microenvironment, and regulate immune function. This paper reviewed the latest research progress on Xuefu Zhuyutang in the treatment of malignant tumors from four aspects: theoretical exploration, clinical studies, mechanisms of action, and pharmacological basis, aiming to provide insights and methods for the clinical diagnosis and treatment of malignant tumors.
9.Neuroprotective Mechanism of Yifei Xuanfei Jiangzhuo Prescription on VaD Rats Based on NF-κB/NLRP3 Signaling Pathway
Bingmao YUAN ; Wei CHEN ; Xiu LAN ; Lingfei JIANG ; Lin WU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(3):88-96
ObjectiveTo investigate the molecular mechanism by which Yifei Xuanfei Jiangzhuo prescription regulates the nuclear factor-κB (NF-κB)/NOD-like receptor protein 3 (NLRP3) signaling pathway to improve neuronal function in vascular dementia (VaD) rats. MethodsA VaD model was established by intermittently clamping the bilateral common carotid arteries (CCA) combined with bilateral vascular occlusion (2-VO). Eighty-four SD rats were randomly divided into a blank group, sham group, model group, piracetam group (0.2 g·kg-1), and low-, medium-, and high-dose Yifei Xuanfei Jiangzhuo prescription groups (6.09, 12.18, and 24.36 g·kg-1). Drug administration began on day 7 after surgery, once daily for 28 consecutive days. Behavioral experiments were used to evaluate learning and spatial memory. Hematoxylin-eosin (HE) staining was applied to observe pathological morphological changes in the CA1 region of the hippocampus. Transmission electron microscopy was used to examine the ultrastructure of hippocampal neurons. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) was used to detect neuronal apoptosis in the CA1 region. Immunohistochemistry was performed to determine the positive expression rate of neuronal nuclear antigen (NeuN). Immunofluorescence single staining was used to assess nuclear expression of NF-κB p65 in brain tissue. Western blot was used to detect the protein expression levels of inhibitor of κB kinase (IKK), NF-κB p65, NLRP3, Caspase-1, apoptosis-associated speck-like protein (ASC), and interleukin-1β (IL-1β). ResultsCompared with the blank group, the model group showed a significant reduction in platform-crossing frequency (P0.01), aggravated hippocampal injury, a significant increase in neuronal apoptosis (P0.05), decreased NeuN positivity in the CA1 region (P0.05), increased nuclear expression of NF-κB p65 (P0.05), and significantly elevated expression of p-IKK, p-NF-κB p65, NLRP3, cleaved Caspase-1, ASC, and cleaved IL-1β (P0.05). Compared with the model group, all drug-treated groups improved learning and spatial memory in VaD rats, alleviated hippocampal pathological injury and neuronal apoptosis, and protected neuronal ultrastructure. Yifei Xuanfei Jiangzhuo prescription at doses of 12.18 and 24.36 g·kg-1 reduced hippocampal expression levels of p-IKK, p-NF-κB p65, NLRP3, Caspase-1, ASC, and cleaved IL-1β in VaD rats (P0.05), showing dose-dependent inhibition of the NF-κB/NLRP3 signaling pathway. ConclusionYifei Xuanfei Jiangzhuo prescription may exert neuroprotective effects by regulating the NF-κB/NLRP3 signaling pathway, thereby reducing neuroinflammation and inhibiting hippocampal neuronal apoptosis.
10.Exploring Mechanism of Yiqi Huoxue Jiedu Formula in Alleviating Immune Cell Exhaustion in Sepsis Based on Transcriptomics and Metabolomics
Rui CHEN ; Qiusha PAN ; Kaiqiang ZHONG ; Shuqi MA ; Wei HUANG ; Jiahua LAI ; Ruifeng ZENG ; Xiaotu XI ; Jun LI
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(3):109-118
ObjectiveTo observe the effects of Yiqi Huoxue Jiedu formula(YHJF) on immune cell exhaustion in the spleen of septic mice and to explore and validate its potential intervention targets. MethodsMice were randomly divided into the sham-operated, model, low-dose YHJF(4.1 g·kg-1), and high-dose YHJF(8.2 g·kg-1) groups. Except for the sham-operated group, a cecal ligation and puncture(CLP) procedure was performed to establish a mouse sepsis model. The treatment groups received oral administration of the corresponding doses, while the sham-operated and model groups received an equal volume of physiological saline. After the intervention, the 7-day survival rate of each group was recorded, and spleen samples were collected 72 h post-intervention, and the spleen index was calculated. Terminal deoxynucleotidyl transferase deoxyuridine triphosphate(dUTP) nick end labeling(TUNEL) staining was used to detect apoptosis in spleen cells. Enzyme-linked immunosorbent assay(ELISA) was performed to measure the levels of interleukin(IL)-4 and IL-10 in the serum. Transcriptomics and metabolomics were used to screen for differentially expressed genes(DEGs) and differential metabolites in the spleen, followed by bioinformatics analysis to identify key targets. Real-time quantitative polymerase chain reaction(Real-time PCR), flow cytometry, and multiplex immunofluorescence were used to verify the expressions of key genes and proteins. ResultsThe high-dose YHJF group significantly improved the 7-day survival rate of septic mice(P0.05). Compared with the sham-operated group, the model group showed a significant increase in apoptosis of spleen cells and a decrease in the spleen index at 72 h post-modeling, with markedly elevated peripheral serum IL-4 and IL-10 levels(P0.01). Compared with the model group, the high-dose YHJF group showed a reduction in apoptosis of spleen cells, an increase in the spleen index, and a significant decrease in peripheral serum IL-4 and IL-10 levels(P0.05). Spleen transcriptomics identified 255 DEGs between groups, potentially serving as intervention targets for YHJF. Gene Ontology(GO) enrichment analysis revealed that DEGs were mainly involved in biological processes such as natural killer(NK) cell-mediated positive immune regulation, cell killing, cytokine production, positive regulation of innate immune cells, and interferon production. Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analysis showed that DEGs were mainly involved in cytokine-cytokine receptor interactions, viral protein interactions with cytokines and cytokine receptors, chemokine signaling pathway, and nuclear transcription factor-κB(NF-κB) signaling pathway. Protein-protein interaction(PPI) network analysis identified CD160, granzyme B(GZMB), and chemokine ligand 4(CCL4) as key targets for YHJF in treating sepsis. Metabolomics identified 46 differential metabolites that were significantly reversed by YHJF intervention, and combined transcriptomics and metabolomics analysis identified 17 differential metabolites closely related to CD160. Pathway enrichment revealed that these metabolites were mainly involved in glycerophospholipid metabolism, arachidonic acid metabolism, glycosylphosphatidylinositol(GPI) anchor biosynthesis, linoleic acid metabolism, and α-linolenic acid metabolism pathways. Verification results showed that, compared with the sham-operated group, the model group exhibited significantly elevated CD160 mRNA expression level in the spleen, along with markedly decreased CCL4 and GZMB mRNA expression, and had a significant increase in CD160 expression on the surface of natural killer T(NKT) cells in the spleen(P0.01). Compared with the model group, the high-dose YHJF group had a significant decrease in CD160 mRNA expression in the spleen, a significant increase in CCL4 and GZMB mRNA expressions. Further flow cytometry and immunofluorescence revealed that compared with the sham-operated group, CD160 expression on the surface of splenic NKT cells in the model group was significantly increased(P0.01), while high-dose YHJF intervention significantly reduced CD160 expression(P0.01). ConclusionYHJF may alleviate NKT cell exhaustion in sepsis by downregulating the expression of the negative co-stimulatory molecule CD160, and this regulatory effect is closely related to fatty acid metabolism pathways. This study provides new insights and targets for further exploration of strengthening vital Qi and detoxifying strategy to improve immune cell exhaustion in acute deficiency syndrome of sepsis.

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