OBJECTIVE: To observe the effects of moxibustion at "Xinshu" (BL15) and "Feishu" (BL13) on myocardial transferrin receptor 1 (TfR1), ferroptosis suppressor protein 1 (FSP1), atrial natriuretic peptide (ANP), and typeⅠcollagen myocardial collagen fibers (CollagenⅠ) in rats with chronic heart failure (CHF), and to explore the mechanism of moxibustion for ameliorating myocardial fibrosis and improving cardiac function in CHF. METHODS: Fifty SD rats were randomly divided into a normal group (n=10) and a modeling group (n=40). The CHF model was established in the modeling group by ligating the left anterior descending coronary artery. After successful modeling, the rats were randomly divided into a model group (n=9), a moxibustion group (n=8), a rapamycin (RAPA) group (n=9), and a moxibustion+RAPA group (n=9). In the moxibustion group, moxibustion was delivered at bilateral "Feishu"(BL13) and "Xinshu" (BL15), 15 min at each point in each intervention, once daily, for 4 consecutive weeks. In the RAPA group, RAPA solution was administered intraperitoneally at a dose of 1 mg/kg, once daily for 4 consecutive weeks. In the moxibustion+RAPA group, RAPA solution was administered intraperitoneally after moxibustion. Ejection fraction (EF) and left ventricular fractional shortening (FS) were measured after modeling and intervention. After intervention, morphology of cardiac muscle was observed using HE staining and Masson's trichrome staining. Total iron content in myocardial tissue was detected using a colorimetric method. Western blot and qPCR were adopted to detect the protein and mRNA expression of TfR1, FSP1, ANP, and CollagenⅠ in myocardial tissue. RESULTS: Compared with the normal group, the EF and FS values decreased (P<0.01); necrosis, edema, degeneration, and arrangement disorder were presented in cardiomyocytes; inflammatory cells were obviously infiltrated, the structure of myocardial fibers was disarranged, the collagen fibers were obviously deposited and fibrosis increased (P<0.01); the total iron content and the protein and mRNA expression of TfR1, ANP, and CollagenⅠ in myocardial tissue were elevated (P<0.01), while the protein and mRNA expression of FSP1 were reduced (P<0.01) in the model group. Compared with the model group, the moxibustion group showed that EF and FS increased (P<0.01); myocardial cell morphology was improved, and myocardial fibrosis was alleviated (P<0.01); the total iron content and the protein and mRNA expression of TfR1, ANP, and CollagenⅠ in myocardial tissue decreased (P<0.01), while the protein and mRNA expression of FSP1 increased (P<0.01, P<0.05). Compared with the model group, the myocardial fibrosis was increased (P<0.05); the total iron content and the protein and mRNA expression of TfR1, ANP, CollagenⅠ in myocardial tissue were increased (P<0.01), while protein and mRNA expression of FSP1 decreased (P<0.01) in the RAPA group. When compared with the RAPA group and the moxibustion + RAPA group, EF and FS were elevated (P<0.01, P<0.05); myocardial cells were improved in morphology, the total iron content and the protein and mRNA expression of TfR1, ANP, and CollagenⅠ in myocardial tissue decreased (P<0.01), while protein and mRNA expression of FSP1 increased (P<0.01) in the moxibustion group. In comparison with the moxibustion + RAPA group, the RAPA group showed the decrease in EF and FS (P<0.01), the worsened myocardial fibrosis (P<0.01), the increase in the total iron content and the protein and mRNA expression of TfR1, ANP, and CollagenⅠ in myocardial tissue (P<0.01), and the decrease in the protein and mRNA expression of FSP1 (P<0.01). CONCLUSION Moxibustion at "Feishu" (BL13) and "Xinshu" (BL15) can slow down the process of myocardial fibrosis and improve cardiac function in CHF rats. The mechanism of moxibustion may be related to inhibiting ferroptosis through regulating autophagy.
Animals ; Rats ; Heart Failure/physiopathology* ; Moxibustion ; Rats, Sprague-Dawley ; Male ; Receptors, Transferrin/genetics* ; Myocardium/metabolism* ; Acupuncture Points ; Humans ; Chronic Disease/therapy* ; Antigens, CD/metabolism*

BACKGROUND: Several studies have demonstrated the occurrence of secondary tumors as a rare but significant complication of chimeric antigen receptor T (CAR-T) cell therapy, underscoring the need for a detailed investigation. Given the limited variety of secondary tumor types reported to date, a comprehensive characterization of the various secondary tumors arising after CAR-T therapy is essential to understand the associated risks and to define the role of the immune microenvironment in malignant transformation. This study aims to characterize the immune microenvironment of a newly identified secondary tumor post-CAR-T therapy, to clarify its pathogenesis and potential therapeutic targets. METHODS: In this study, the bone marrow (BM) samples were collected by aspiration from the primary and secondary tumors before and after CD19 CAR-T treatment. The CD45 + BM cells were enriched with human CD45 microbeads. The CD45 + cells were then sent for 10× genomics single-cell RNA sequencing (scRNA-seq) to identify cell populations. The Cell Ranger pipeline and CellChat were used for detailed analysis. RESULTS: In this study, a rare type of secondary chronic myelomonocytic leukemia (CMML) were reported in a patient with diffuse large B-cell lymphoma (DLBCL) who had previously received CD19 CAR-T therapy. The scRNA-seq analysis revealed increased inflammatory cytokines, chemokines, and an immunosuppressive state of monocytes/macrophages, which may impair cytotoxic activity in both T and natural killer (NK) cells in secondary CMML before treatment. In contrast, their cytotoxicity was restored in secondary CMML after treatment. CONCLUSIONS This finding delineates a previously unrecognized type of secondary tumor, CMML, after CAR-T therapy and provide a framework for defining the immune microenvironment of secondary tumor occurrence after CAR-T therapy. In addition, the results provide a rationale for targeting macrophages to improve treatment strategies for CMML treatment.
Humans ; Lymphoma, Large B-Cell, Diffuse/therapy* ; Tumor Microenvironment/genetics* ; Antigens, CD19/metabolism* ; Leukemia, Myelomonocytic, Chronic/genetics* ; Immunotherapy, Adoptive/adverse effects* ; Male ; Single-Cell Analysis/methods* ; Female ; Sequence Analysis, RNA/methods* ; Receptors, Chimeric Antigen ; Middle Aged

Neoantigens exhibit high immunogenic potential and confer a uniqueness to tumor cells, making them ideal targets for personalized cancer immunotherapy. Neoantigens originate from tumor-specific genetic alterations, abnormal viral infections, or other biological mechanisms, including atypical RNA splicing events and post-translational modifications (PTMs). These neoantigens are recognized as foreign by the immune system, eliciting an immune response that largely bypasses conventional mechanisms of central and peripheral tolerance. Advances in next-generation sequencing (NGS), mass spectrometry (MS), and artificial intelligence (AI) have greatly expedited the rapid detection and forecasting of neoantigens, markedly propelling the development of diverse immunotherapeutic strategies, including cancer vaccines, adoptive cell therapy, and antibody treatment. In this review, we comprehensively explore the discovery and characterization of neoantigens and their clinical use within promising immunotherapeutic frameworks. Additionally, we address the current landscape of neoantigen research, the intrinsic challenges of the field, and potential pathways for clinical application in cancer treatment.
Humans ; Neoplasms/therapy* ; Precision Medicine/methods* ; Immunotherapy/methods* ; Antigens, Neoplasm/genetics* ; Cancer Vaccines/immunology* ; High-Throughput Nucleotide Sequencing

The study aimed to investigate the effect of the CD200R1 gene deletion on microglia activation and nigrostriatal dopamine neuron loss in the Parkinson's disease (PD) process. The CRISPR-Cas9 technology was applied to construct the CD200R1^-/- mice. The primary microglia cells of wild-type and CD200R1^-/- mice were cultured and treated with bacterial lipopolysaccharide (LPS). Microglia phagocytosis level was assessed by a fluorescent microsphere phagocytosis assay. PD mouse model was prepared by nigral stereotaxic injection of recombinant adeno-associated virus vector carrying human α-synuclein (α-syn). The changes in the motor behavior of the mice with both genotypes were evaluated by cylinder test, open field test, and rotarod test. Immunohistochemical staining was used to assess the loss of dopamine neurons in substantia nigra. Immunofluorescence staining was used to detect the expression level of CD68 (a key molecule involved in phagocytosis) in microglia. The results showed that CD200R1 deletion markedly enhanced LPS-induced phagocytosis in vitro by the microglial cells. In the mouse model of PD, CD200R1 deletion exacerbated motor behavior impairment and dopamine neuron loss in substantia nigra. Fluorescence intensity analysis results revealed a significant increase in CD68 expression in microglia located in the substantia nigra of CD200R1^-/- mice. The above results suggest that CD200R1 deletion may further activates microglia by promoting microglial phagocytosis, leading to increased loss of the nigrostriatal dopamine neurons in the PD model mice. Therefore, targeting CD200R1 could potentially serve as a novel therapeutic target for the treatment of early-stage PD.
Animals ; Microglia/physiology* ; Mice ; Phagocytosis ; Parkinson Disease/genetics* ; Disease Models, Animal ; Receptors, Cell Surface/physiology* ; Dopaminergic Neurons/pathology* ; Antigens, CD/metabolism* ; Gene Deletion ; Substantia Nigra ; Mice, Inbred C57BL ; Mice, Knockout ; Cells, Cultured ; Male ; alpha-Synuclein ; CD68 Molecule ; Orexin Receptors

The current study aimed to clarify the roles of apolipoprotein A5 (ApoA5) and milk fat globule-epidermal growth factor 8 (Mfge8) in regulating myocardial lipid deposition and the regulatory relationship between them. The serum levels of ApoA5 and Mfge8 in obese and healthy people were compared, and the obesity mouse model induced by the high-fat diet (HFD) was established. In addition, primary cardiomyocytes were purified and identified from the hearts of suckling mice. The 0.8 mmol/L sodium palmitate treatment was used to establish the lipid deposition cardiomyocyte model in vitro. ApoA5-overexpressing adenovirus was used to observe its effects on cardiac function and lipids. The expressions of the fatty acid uptake-related molecules and Mfge8 on transcription or translation levels were detected. Co-immunoprecipitation was used to verify the interaction between ApoA5 and Mfge8 proteins. Immunofluorescence was used to observe the co-localization of Mfge8 protein with ApoA5 or lysosome-associated membrane protein 2 (LAMP2). Recombinant rMfge8 was added to cardiomyocytes to investigate the regulatory mechanism of ApoA5 on Mfge8. The results showed that participants in the simple obesity group had a significant decrease in serum ApoA5 levels (P < 0.05) and a significant increase in Mfge8 levels (P < 0.05) in comparison with the healthy control group. The adenovirus treatment successfully overexpressed ApoA5 in HFD-fed obese mice and palmitic acid-induced lipid deposition cardiomyocytes, respectively. ApoA5 reduced the weight of HFD-fed obese mice (P < 0.05), shortened left ventricular isovolumic relaxation time (IVRT), increased left ventricular ejection fraction (LVEF), and significantly reduced plasma levels of triglycerides (TG) and cholesterol (CHOL) (P < 0.05). In myocardial tissue and cardiomyocytes, the overexpression of ApoA5 significantly reduced the deposition of TG (P < 0.05), transcription of fatty acid translocase (FAT/CD36) (P < 0.05), fatty acid-binding protein (FABP) (P < 0.05), and fatty acid transport protein (FATP) (P < 0.05), and protein expression of Mfge8 (P < 0.05), while the transcription levels of Mfge8 were not significantly altered (P > 0.05). In vitro, the Mfge8 protein was captured using ApoA5 as bait protein, indicating a direct interaction between them. Overexpression of ApoA5 led to an increase in co-localization of Mfge8 with ApoA5 or LAMP2 in cardiomyocytes under lipid deposition status. On this basis, exogenous added recombinant rMfge8 counteracted the improvement of lipid deposition in cardiomyocytes by ApoA5. The above results indicate that the overexpression of ApoA5 can reduce fatty acid uptake in myocardial cells under lipid deposition status by regulating the content and cellular localization of Mfge8 protein, thereby significantly reducing myocardial lipid deposition and improving cardiac diastolic and systolic function.
Animals ; Humans ; Mice ; Myocytes, Cardiac/metabolism* ; Obesity/physiopathology* ; Male ; Apolipoprotein A-V/blood* ; Lipid Metabolism/physiology* ; Milk Proteins/blood* ; Myocardium/metabolism* ; Diet, High-Fat ; Antigens, Surface/physiology* ; Mice, Inbred C57BL ; Cells, Cultured ; Female

Objective This study aims to investigate and analyze the distribution of MN blood type among ethnic minorities in China. Methods Through a systematic retrieval of the 981 literature related to MN blood group distribution, 120 literature, meeting the criteria of this study, with complete data were selected. The literature covers 49 ethnic minorities. SPSS 26 statistical software was used to analyze the data. Results The results showed that among the 49 ethnic minorities in China, the phenotype distribution of MN blood type was MN>MM>NN, with proportions of 42.54%, 41.86%, and 15.06% respectively. The gene frequency for MN blood type exhibited a trend of m>n, with a gene frequency of m being 0.6313 and n being 0.3687. Cluster analysis divided the Chinese ethnic minorities into three groups based on the gene frequency for m, showing the characteristics of Group I>Group II>Group III. Conclusion The MN blood type characteristics in Chinese ethnic minorities show a higher frequency of the M gene compared to the N gene. The frequency of the M gene is higher in southern ethnic minorities than in northern ones. There are significant differences between southwestern ethnic minorities and the Han nationality, but no differences with long-term mixed/settled Han populations.
Humans ; China/ethnology* ; Minority Groups ; Ethnicity/genetics* ; Gene Frequency ; Asian People/genetics* ; Blood Group Antigens/genetics*

Programmed death 1 (PD-1) and its ligand (PD-L1) serve as crucial targets in cancer immunotherapy, and their inhibitors have significantly improved the prognosis of many patients with malignant tumors. However, the issues of drug resistance and limited overall response rate associated with monotherapy remain prevalent. As a new generation of immune checkpoints, lymphocyte activation gene 3 (LAG-3) synergistically enhances the suppression of T cells alongside PD-1 in various cancers. Combining the blockade of both PD-1 and LAG-3 yields stronger anti-tumor immune effects compared to blocking either target alone, thereby reversing the immunosuppressive state of the tumor microenvironment and reducing the occurrence of resistance. This review covers the structural characteristics of LAG-3 and unveils its specific interactions with PD-1 across multiple cancers, providing a novel reference for overcoming the limitations of single-agent therapy.
Humans ; Neoplasms/immunology* ; Immunotherapy/methods* ; Programmed Cell Death 1 Receptor/metabolism* ; Lymphocyte Activation Gene 3 Protein ; Antigens, CD/metabolism* ; Animals ; Tumor Microenvironment/immunology* ; Immune Checkpoint Inhibitors/therapeutic use*

Objective To investigate the effects of LBL-007, a novel fully human lymphocyte activation gene 3 (LAG-3) monoclonal antibody, in combination with programmed cell death protein 1 (PD-1) antibody, on the invasion, migration and proliferation of tumor cells, and to elucidate the underlying mechanisms. Methods Human lymphocyte cells Jurkat were co-cultured with A549 and MGC803 tumor cell lines and treated with the isotype control antibody human IgG, LBL-007, anti-PD-1 antibody BE0188, or tumor necrosis factor-alpha (TNF-α, the NF-κB signaling pathway agonist). Tumor cell proliferation was assessed using a colony formation assay; invasion was measured by Transwell^TM assay; migration was evaluated using a wound healing assay. Western blotting was employed to determine the expression levels of NF-κB pathway-related proteins: IκB inhibitor kinase alpha (Ikkα), phosphorylated Ikkα (p-IKKα), NF-κB subunit p65, phosphorylated p65 (p-p65), NF-κB Inhibitor Alpha (IκBα), phosphorylated IκBα (p-IκBα), matrix metalloproteinase 9 (MMP9), and MMP2. Results Compared with the control and IgG isotype groups, LBL-007 and BE0188 significantly reduced tumor cell proliferation, invasion, and migration. They also decreased the phosphorylation of p-IKKα, p-p65 and p-IκBα, and the expression of MMP9 and MMP2 of tumor cells in the co-culture system. The combined treatment of LBL-007 and BE0188 enhanced inhibitory effects. Treatment with the NF-κB signaling pathway agonist TNF-α reversed the suppressive effects of LBL-007 and BE0188 on tumor cell proliferation, invasion, migration, and NF-κB signaling. Conclusion LBL-007 and anti-PD-1 antibody synergistically inhibit the invasion, migration, and proliferation of A549 and MGC803 tumor cells by blocking the NF-κB signaling pathway.
Humans ; Cell Proliferation/drug effects* ; Cell Movement/drug effects* ; Signal Transduction/drug effects* ; NF-kappa B/metabolism* ; Neoplasm Invasiveness ; Antibodies, Monoclonal/pharmacology* ; Programmed Cell Death 1 Receptor/antagonists & inhibitors* ; Cell Line, Tumor ; Antigens, CD/immunology* ; Lymphocyte Activation Gene 3 Protein ; A549 Cells ; I-kappa B Kinase/metabolism* ; Jurkat Cells ; Matrix Metalloproteinase 9/metabolism*

Objective To prepare monoclonal antibodies against bovine CD4 and identify their basic biological characteristics. Methods Recombinant bovine CD4 (rHis-BoCD4 and rGST-BoCD4) was successfully expressed and purified by constructing a prokaryotic plasmid of bovine CD4 gene. The bovine CD4 monoclonal antibody was produced using hybridoma technology. The subtype and potency of the monoclonal antibody were identified and analyzed by ELISA, while specificity was analyzed through indirect immunofluorescence assay (IFA) and Western-blot. Results Four hybridoma cell lines, namely, 1H4, 6A10, 3F9 and 4G10, stably secreting monoclonal antibodies against BoCD4 were successfully obtained. The subclasses of the monoclonal antibodies subclass 6A10 was IgG2b and the rest of the monoclonal antibodies were of IgM type. Western-blot results showed that the four anti-bovine CD4 mAb strains were able to specifically bind to the bovine CD4 protein expressed in vitro. Indirect immunofluorescence assay showed that four monoclonal antibodies were able to specifically recognize the natural bovine CD4 protein. Flow cytometry assay showed that 3F9 was best to recognize bovine natural CD4 molecules. Conclusion Four monoclonal antibody strains with high specificity to natural bovine CD4 protein were successfully prepared, which lays the foundation for the subsequent studies on the function of bovine CD4 and diagnosis and treatment of bovine T-lymphocyte diseases.
Animals ; Antibodies, Monoclonal/isolation & purification* ; Cattle ; CD4 Antigens/genetics* ; Hybridomas/immunology* ; Antibody Specificity/immunology* ; Mice ; Mice, Inbred BALB C ; Enzyme-Linked Immunosorbent Assay ; Fluorescent Antibody Technique, Indirect

Objective To generate and characterize natural killer cell (NK cell)-conditional Cd226 gene knockout mice using Cre-loxP technology, and to explore the role of CD226 on NK cells in alleviating intestinal inflammation in a murine model of ulcerative colitis (UC). Methods NK cell-conditional Cd226 gene knockout mice were generated by crossing loxP-flanked Cd226 mice with Ncr1-Cre mice via the Cre-loxP system. Polymerase chain reaction (PCR) and agarose gel electrophoresis were used for genotyping. A UC model was established by dextran sulfate sodium (DSS) induction. Flow cytometry was performed to analyze CD226 expression levels on NK cells and the infiltration of related immune cells in colon tissues. Hematoxylin-eosin (HE) staining was performed to assess the degree of colonic inflammation. Results DNA gel electrophoresis and flow cytometry confirmed the successful generation of NK cell-specific Cd226 knockout mice. After conditional knockout of Cd226 in NK cells, inflammation in the UC mouse model was alleviated. Flow cytometry results showed a reduced proportion of NK cells in peripheral blood and the colon lamina propria, while HE staining demonstrated attenuated inflammatory responses. Conclusion Specific knockout of Cd226 in NK cells mitigates intestinal inflammation in UC mice by reducing NK cell numbers and inhibiting their pro-inflammatory functions.
Animals ; Colitis, Ulcerative/pathology* ; Killer Cells, Natural/metabolism* ; Mice, Knockout ; T Lineage-Specific Activation Antigen 1 ; Antigens, Differentiation, T-Lymphocyte/genetics* ; Mice ; Disease Models, Animal ; Mice, Inbred C57BL ; Male