Metabolic diseases have seen a steady increase in incidence in recent years, becoming one of the main causes of sub-health status globally. Neutrophil extracellular traps(NETs) are reticular complexes containing DNA, which trap foreign microorganisms or induce an immune response. Current research indicates that NETs are widely active in various metabolic diseases and can cause severe damage to the body through multiple mechanisms, including promoting blood glucose elevation, damaging vascular endothelial cells, forming vascular embolisms, triggering intense inflammation, and promoting lipid accumulation. Therefore, intervening in NETs is an important approach to treating metabolic diseases. Research has shown a close relationship between the theory of spleen heat-turbid toxin theory and metabolic diseases-NETs mechanism. The basic pathogenesis include the internal accumulation of phlegm-dampness, qi stagnation and blood stasis, internal accumulation of dampness-heat, phlegm and blood stasis, and flourishing toxic heat. Various Chinese herbal medicines with the functions of dispelling dampness, resolving phlegm, promoting blood circulation to remove blood stasis, and clearing heat and toxins, along with their extracts and compound prescriptions, can treat metabolic diseases by regulating NETs and delaying disease progression. This paper systematically outlined the formation mechanisms of NETs, their connection to metabolic diseases, the theoretical basis in TCM, their roles in numerous metabolic diseases, and the current research status of TCM in regulating NETs to prevent and control metabolic diseases, aiming to provide effective reference ideas for developing therapeutic strategies for metabolic diseases.
Humans ; Extracellular Traps/metabolism* ; Metabolic Diseases/metabolism* ; Drugs, Chinese Herbal/therapeutic use* ; Animals ; Neutrophils/metabolism* ; Medicine, Chinese Traditional

Asthma is a chronic inflammatory disease involving multiple inflammatory cells and cytokines. Its pathogenesis is complex, involving various cells and cytokines. Traditional Chinese medicine(TCM) theory suggests that the pathogenesis of asthma is closely related to the dysfunction of internal organs such as the lungs, spleen, and kidneys. In contrast, modern immunological studies have revealed the central role of T helper 1(Th1)/T helper 2(Th2) and T helper 17(Th17)/regulatory T(Treg) cellular immune imbalance in the pathogenesis of asthma. Th1/Th2 imbalance is manifested as hyperfunction of Th2 cells, which promotes the synthesis of immunoglobulin E(IgE) and the activation of eosinophil granulocytes, leading to airway hyperresponsiveness and inflammation.Meanwhile, Th17/Treg imbalance exacerbates the inflammatory response in the airways, further contributing to asthma pathology.Currently, therapeutic strategies for asthma are actively exploring potential targets for regulating the balance of Th1/Th2 and Th17/Treg immune responses. These targets include cytokines, transcription factors, key proteins, and non-coding RNAs. Precisely regulating the expression and function of these targets can effectively modulate the activation and differentiation of immune cells. In recent years,traditional Chinese medicine active ingredients have shown unique potential and prospects in the field of asthma treatment. Based on this, the present study systematically summarizes the efficacy and specific mechanisms of TCM active ingredients in treating asthma by regulating Th1/Th2 and Th17/Treg immune balance through literature review and analysis. These active ingredients, including flavonoids, terpenoids, polysaccharides, alkaloids, and phenolic acids, exert their effects through various mechanisms, such as inhibiting the activation of inflammatory cells, reducing the release of cytokines, and promoting the normal differentiation of immune cells. This study aims to provide a solid foundation for the widespread application and in-depth development of TCM in asthma treatment and to offer new ideas for clinical research and drug development of asthma.
Asthma/genetics* ; Humans ; Drugs, Chinese Herbal/chemistry* ; Th2 Cells/drug effects* ; Th17 Cells/drug effects* ; T-Lymphocytes, Regulatory/drug effects* ; Th1 Cells/drug effects* ; Animals ; Cytokines/immunology* ; Medicine, Chinese Traditional

OBJECTIVES: To investigate the therapeutic effects of cimifugin on Crohn's disease (CD)-like colitis in mice and its possible mechanism. METHODS: Thirty adult male C57BL/6 mice were randomized equally into control group, 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced CD-like colitis model group, and cimifugin treatment (daily gavage at 12.5 mg/kg) group. The therapeutic effect of cimifugin was evaluated by observing changes in body weight, disease activity index (DAI) scores, colon length, histopathological inflammation scores, and inflammatory cytokine levels in the colonic mucosa. Intestinal barrier integrity in the mice was assessed using immunofluorescence assay and Western blotting for claudin-1 and ZO-1; T-helper (Th) cell subset ratios in the mesenteric lymph nodes were analyzed with flow cytometry. Network pharmacology, KEGG enrichment analysis and molecular docking were used to predict the targets of cimifugin and analyze the key pathways and cimifugin-MAPK protein interactions, which were validated by Western blotting in the mouse models. RESULTS: In mice with TNBS-induced colitis, cimifugin treatment significantly attenuated body weight loss and colon shortening, lowered DAI and histopathological scores, decreased IFN-γ and IL-17 levels, and increased IL-4 and IL-10 levels in the colonic mucosa. Cimifugin treatment also significantly improved TNBS-induced claudin-1 dislocation and reduction of goblet cells, upregulated claudin-1 and ZO-1 expressions, reduced Th1 and Th17 cell percentages, and increased Th2 and Treg cell percentages in the colonic mucosa of the mice. KEGG analysis suggested a possible connection between the effect of cimifugin and MAPK signaling, and molecular docking showed strong binding affinity between cimifugin and MAPK core proteins. Western blotting demonstrated significantly decreased phosphorylation levels of JNK, ERK, and p38 in the colonic mucosa of cimifugin-treated mouse models. CONCLUSIONS Cimifugin alleviates TNBS-induced CD-like colitis by repairing intestinal barrier damage and restoring Th1/Th2 and Th17/Treg balance via suppressing MAPK pathway activation.
Animals ; Mice, Inbred C57BL ; Male ; Mice ; Crohn Disease/immunology* ; Colitis/immunology* ; MAP Kinase Signaling System/drug effects* ; Trinitrobenzenesulfonic Acid ; T-Lymphocytes, Helper-Inducer/drug effects* ; Intestinal Mucosa ; Disease Models, Animal

OBJECTIVES: To investigate the characteristics of immune cell infiltration in tumor samples from Chinese patients with clear cell renal cell carcinoma (ccRCC) and the correlation of immune cell infiltration with tumor stage and response to immunotherapy. METHODS: Tumor samples and clinicopathological data were collected from 154 ccRCC patients treated in Nanfang Hospital, Southern Medical University from October, 2020 to October, 2023. The immune cell types infiltrating the tumor tissues were identified using immunohistochemistry and immunofluorescence staining, and their correlations with the patients' clinicopathological characteristics were analyzed. Patient-derived tumor tissue fragment models (PDTF) models, constructed using tumor tissues from 22 patients, were treated with PD-1 monoclonal antibody, and T cell activation was detected using flow cytometry to assess the patients' responses to immunotherapy. RESULTS: In Chinese ccRCC patients included in this study, CD8⁺ T cells, CD4⁺ T cells, and CD3⁺ T cells were the most abundant in the tumor tissues. Higher infiltration levels of CD3⁺ T cells (P=0.004), PD-1⁺ T cells (P=0.020), CD68⁺ T cells (P=0.049), CD79⁺ T cells (P=0.049), and Tryptase⁺ cells (P=0.049) were all positively correlated with a larger tumor size (≥5 cm). A higher infiltration level of CD4⁺ T cells was associated with a lower tumor stage. Patients with higher International Society of Urological Pathology (ISUP) grades had higher infiltration levels of CD3⁺ T cells (P=0.023), CD8⁺ T cells (P=0.045), PD-1⁺ T cells (P=0.014), CD20⁺ B cells (P=0.020) and CD79⁺ B cells (P=0.049), and lower levels of Tryptase⁺ cells (P=0.001). Patients with abundant infiltrating immune cells tended to have better responses to immunotherapy. CONCLUSIONS The infiltrating immune cells are heterogeneous in Chinese ccRCC patients, and immune cell infiltration characteristics are closely correlated with clinicopathological parameters of the patients.
Humans ; Carcinoma, Renal Cell/pathology* ; Kidney Neoplasms/pathology* ; Immunotherapy ; Male ; Lymphocytes, Tumor-Infiltrating/immunology* ; Female ; Middle Aged ; CD8-Positive T-Lymphocytes/immunology* ; Aged ; T-Lymphocytes/immunology* ; Programmed Cell Death 1 Receptor/immunology* ; Adult ; CD4-Positive T-Lymphocytes/immunology* ; Neoplasm Staging

OBJECTIVES: To investigate the inhibitory effect of pirfenidone (PFD) on growth of bladder cancer xenograft and its regulatory effect on Treg cells in tumor-bearing mice. METHODS: Thirty-two C57BL/6 mice bearing ectopic bladder tumors were randomized into control and PFD groups (n=16). In PFD group, PFD was administered orally at the daily dose of 500 mg/kg, and tumor growth and survival of the mice were monitored. After treatment for 21 days, the tumors and vital organs were harvested for analysis. Immunohistochemistry was used to assess CD3, CD4, CD8, and FOXP3 expressions in the tumors. Flow cytometry and RT-qPCR were used to analyze the percentage of CD4⁺CD25⁺FOXP3⁺ Treg cells and IL-2, IL-10, and IL-35 expressions in the tumors and spleens; organ damage of the mice was examined with HE staining. RESULTS: Compared with the control group, the PFD-treated mice exhibited significantly lower tumor growth rate with smaller tumor volumes at day 21, along with improved survival at day 28. Immunohistochemistry revealed no significant differences in the infiltration of CD3⁺ and CD8⁺ cells between the two groups, but the percentages of CD4⁺ and FOXP3⁺ cells were significantly lower in the tumors of PFD-treated mice. Flow cytometric analysis confirmed a decrease in CD4⁺CD25⁺FOXP3⁺ Treg cells in the tumors from PFD-treated mice, which also had reduced expression levels of IL-2, IL-10 and IL-35 mRNAs in the tumors. No significant differences were found in Treg cell populations or cytokine expressions in the spleen tissues between the two groups. HE staining showed obvious organ damage in neither of the groups. CONCLUSIONS PFD inhibits bladder cancer growth and enhances survival of tumor-bearing mice possibly by suppressing Treg cells in the tumor microenvironment.
Animals ; Urinary Bladder Neoplasms/drug therapy* ; Mice ; T-Lymphocytes, Regulatory/metabolism* ; Mice, Inbred C57BL ; Interleukins/metabolism* ; Interleukin-10/metabolism* ; Cell Line, Tumor ; Interleukin-2/metabolism* ; Xenograft Model Antitumor Assays ; Female

OBJECTIVES: To identify immunosuppressive neutrophil subsets in patients with prostate cancer (PCa) and construct a risk prediction model for prognosis and immunotherapy response of the patients based on these neutrophil subsets. METHODS: Single-cell and transcriptome data from PCa patients were collected from the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA). Neutrophil subsets in PCa were identified through unsupervised clustering, and their biological functions and effects on immune regulation were analyzed by functional enrichment, cell interaction, and pseudo-time series analyses. Lasso-Cox regression was utilized to construct a prognostic risk model based on the immunosuppressive neutrophil subsets, and survival analysis and ROC curve analysis were used to compare the prognosis of PCa patients with high and low risks stratified using this model. The relationship of the prognostic risk model with PCa immune infiltration and immune response was evaluated using CIBERSORT and TIDE scores. RESULTS: PCa tissues showed a significantly greater proportion of infiltrating neutrophils than the adjacent normal tissues (P<0.05). PCa-associated neutrophils could be clustered into two independent cell subsets: Neu_1 and Neu_2. Neu_2 cells exhibited highly enriched immunoregulatory functions and were highly differentiated and mature, with upregulated immunosuppressive cytokines such as TGFB1, ITGB2, and LGALS3. Based on the genetic characteristics of Neu_2 cell subsets, the prognostic risk model was constructed. The patients in the high-risk group identified by the model had a shorter biochemical recurrence time (P<0.05) and a higher proportion of Tregs and M2-TAMs cell infiltration (P<0.05) with a higher risk of immune rejection and poorer immune response scores. CONCLUSIONS PCa-associated neutrophils are highly heterogeneous. The prognostic risk model constructed based on the immunosuppressive neutrophil Neu_2 subset can effectively predict both the survival outcomes and immune response of PCa patients.
Humans ; Male ; Prostatic Neoplasms/diagnosis* ; Prognosis ; Neutrophils/immunology* ; Immunotherapy

OBJECTIVES: To investigate the therapeutic mechanism of 2,6-dimethoxy-1,4-benzoquinone (DMQ) for alleviating dextran sulfate sodium (DSS)-induced ulcerative colitis (UC) in mice. METHODS: Eighteen male C57BL/6J mice were equally randomized into control group, DSS group and DMQ treatment group. In DSS and DMQ groups, the mice were treated with DSS in drinking water to induce UC, and received intraperitoneal injections of sterile PBS or DMQ (20 mg/kg) during modeling. The changes in body weight, disease activity index (DAI), colon length, spleen weight, and colon histological scores of the mice were examined, and the percentages of Th17 and IFN-γ⁺ CD8⁺ T cells in the mesenteric lymph nodes and spleen were analyzed using flow cytometry. The expressions of tight junction proteins (Occludin and ZO-1), proteins associated with inflammasome activation (caspase-1 and p20), IL-1β and TNF-α in the colon tissues were detected using Western blotting or ELISA. In the cell experiment, mouse bone marrow-derived macrophages (BMDMs) primed with lipopolysaccharide (LPS) were treated with DMQ, followed by stmulation with nigericin to activate the classical NLRP3 inflammasome pathway. In cultured human peripheral blood mononuclear cells (PBMCs) treated with either LPS alone or LPS plus nigericin, the effects of DMQ on inflammasome activation, pyroptosis, and cytokine release were evaluated via Western blotting, ELISA, and flow cytometry. RESULTS: In DSS-treated mice, DMQ treatment significantly alleviated DSS-induced body weight loss, colon shortening, spleen enlargement, and colon inflammation. The DMQ-treated mice showed significantly reduced percentages of Th17 cells and IFN-γ⁺ CD8⁺ T cells in the mesenteric lymph nodes and spleen, with increased occludin and ZO-1 expressions and decreased caspase-1 expression in the colon tissue. DMQ obviously inhibited classical NLRP3 inflammasome activation in mouse BMDMs and both the classical and alternative pathways of NLRP3 activation in human PBMCs, causing also suppression of caspase-1-dependent pyroptosis. CONCLUSIONS DMQ ameliorates DSS-induced UC in mice by inhibiting NLRP3 inflammasome activation.
Animals ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Mice, Inbred C57BL ; Colitis, Ulcerative/metabolism* ; Dextran Sulfate/adverse effects* ; Male ; Inflammasomes/metabolism* ; Mice ; Benzoquinones/therapeutic use* ; Th17 Cells ; Caspase 1/metabolism*

Infiltration and activation of peripheral immune cells are critical in the progression of multiple sclerosis and its experimental animal model, experimental autoimmune encephalomyelitis (EAE). This study investigates the role of high mobility group box 1 (HMGB1) in oligodendrocyte precursor cells (OPCs) in modulating pathogenic T cells infiltrating the central nervous system through the blood-brain barrier (BBB) by using OPC-specific HMGB1 knockout (KO) mice. We found that HMGB1 released from OPCs promotes BBB disruption, subsequently allowing increased immune cell infiltration. The migration of CD4+ T cells isolated from EAE-induced mice was enhanced when co-cultured with OPCs compared to oligodendrocytes (OLs). OPC-specific HMGB1 KO mice exhibited lower BBB permeability and reduced immune cell infiltration into the CNS, leading to less damage to the myelin sheath and mitigated EAE progression. CD4+ T cell migration was also reduced when co-cultured with HMGB1 knock-out OPCs. Our findings reveal that HMGB1 secretion from OPCs is crucial for regulating immune cell infiltration and provides insights into the immunomodulatory function of OPCs in autoimmune diseases.
Animals ; Encephalomyelitis, Autoimmune, Experimental/metabolism* ; HMGB1 Protein/deficiency* ; Mice, Knockout ; Oligodendrocyte Precursor Cells/immunology* ; Mice, Inbred C57BL ; CD4-Positive T-Lymphocytes/immunology* ; Cell Movement ; Blood-Brain Barrier/immunology* ; Mice ; Myelin Sheath/pathology* ; Disease Models, Animal ; Coculture Techniques ; Oligodendroglia/metabolism* ; Female ; Cells, Cultured

The increasing aging population and aging-associated diseases have become a global issue for decades. People over 65 show an increased prevalence and greater severity of periodontitis, which poses threats to overall health. Studies have demonstrated a significant association between aging and the dysfunction of neutrophils, critical cells in the early stages of periodontitis, and their crosstalk with macrophages and T and B lymphocytes to establish the periodontal lesion. Neutrophils differentiate and mature in the bone marrow before entering the circulation; during an infection, they are recruited to infected tissues guided by the signal from chemokines and cytokines to eliminate invading pathogens. Neutrophils are crucial in maintaining a balanced response between host and microbes to prevent periodontal diseases in periodontal tissues. The impacts of aging on neutrophils' chemotaxis, anti-microbial function, cell activation, and lifespan result in impaired neutrophil functions and excessive neutrophil activation, which could influence periodontitis course. We summarize the roles of neutrophils in periodontal diseases and the aging-related impacts on neutrophil functional responses. We also explore the underlying mechanisms that can contribute to periodontitis manifestation in aging. This review could help us better understand the pathogenesis of periodontitis, which could offer novel therapeutic targets for periodontitis.
Humans ; Neutrophils/immunology* ; Periodontitis/immunology* ; Aging/physiology*

Microwave thermochemotherapy (MTC) has been applied to treat lip squamous cell carcinoma (LSCC), but a deeper understanding of its therapeutic mechanisms and molecular biology is needed. To address this, we used single-cell transcriptomics (scRNA-seq) and spatial transcriptomics (ST) to highlight the pivotal role of tumor-associated neutrophils (TANs) among tumor-infiltrating immune cells and their therapeutic response to MTC. MNDA⁺ TANs with anti-tumor activity (N1-phenotype) are found to be abundantly infiltrated by MTC with benefit of increased blood perfusion, and these TANs are characterized by enhanced cytotoxicity, ameliorated hypoxia, and upregulated IL1B, activating T&NK cells and fibroblasts via IL1B-IL1R. In this highly anti-tumor immunogenic and hypoxia-reversed microenvironment under MTC, fibroblasts accumulated in the tumor front (TF) can recruit N1-TANs via CXCL2-CXCR2 and clear N2-TANs (pro-tumor phenotype) via CXCL12-CXCR4, which results in the aggregation of N1-TANs and extracellular matrix (ECM) deposition. In addition, we construct an N1-TANs marker, MX2, which positively correlates with better prognosis in LSCC patients, and employ deep learning techniques to predict expression of MX2 from hematoxylin-eosin (H&E)-stained images so as to conveniently guide decision making in clinical practice. Collectively, our findings demonstrate that the N1-TANs/fibroblasts defense wall formed in response to MTC effectively combat LSCC.
Humans ; Neutrophils/metabolism* ; Single-Cell Analysis ; Lip Neoplasms/genetics* ; Hyperthermia, Induced/methods* ; Microwaves/therapeutic use* ; Transcriptome ; Carcinoma, Squamous Cell/immunology* ; Tumor Microenvironment