RESUMO
The Gasdermin protein is a membrane disruptor that can mediate immunogenic pyroptosis and elicit anti-tumor immune function. However, cancer cells downregulate Gasdermin and develop membrane repair mechanisms to resist pyroptosis. Therefore, an artificial membrane disruptor (AMD) that can directly mediate membrane rupture in pyroptosis-deficient cells and induce antitumor immune responses in a controllable manner will be valuable in preclinical and clinical research. A micron-scale Ce6-based AMD that can directly induce plasma membrane rupture (PMR) in gasdermin-deficient tumor cells is established. Micron-scale AMDs localize Ce6 specifically to the plasma membrane without labeling other organelles. Compared to free Ce6 molecules, the use of AMDs results in a higher degree of specificity for the plasma membrane. Due to this specificity, AMDs mediate fast and irreversible PMR under 660 nm red light. Furthermore, the AMDs are capable of inducing programmed cell death and lytic cell death in a catalytic manner, demonstrating that the amount of Ce6 used by AMDs is only one-fifth of that used by Ce6 alone when inducing 80% of cancer cell death. In vivo, the AMDs show specificity for tumor targeting and penetration, suggesting that light-driven programmed cell death is specific to tumors. AMDs are applied to antitumor therapy in gasdermin-deficient tumors, resulting in efficient tumor elimination with minimal damage to major organs when combined with anti-PD-1 therapy. Tumor regression is correlated with PMR-mediated inflammation and T-cell-based immune responses. This study provides new insights for designing bioinspired membrane disruptors for PMR and mediating anti-tumor immunotherapy. Additionally, AMD is a dependable tool for examining the immunogenicity of PMR both in vitro and in vivo.
Assuntos
Membrana Celular , Animais , Camundongos , Membrana Celular/metabolismo , Humanos , Modelos Animais de Doenças , Linhagem Celular Tumoral , Neoplasias/imunologia , Piroptose/imunologia , Proteínas de Ligação a Fosfato/genética , Proteínas de Ligação a Fosfato/metabolismoRESUMO
Gastric cancer (GC) has high rates of morbidity and mortality, and this phenomenon is particularly evident in coastal regions where local dietary habits favor the consumption of pickled foods such as salted fish and vegetables. In addition, the diagnosis rate of GC remains low due to the lack of diagnostic serum biomarkers. Therefore, in this study, we aimed to identify potential serum GC biomarkers for use in clinical practice. To identify candidate biomarkers of GC, 88 serum samples were first screened using a high-throughput protein microarray to measure the levels of 640 proteins. Then, 333 samples were used to validate the potential biomarkers using a custom antibody chip. ELISA, western blot, and immunohistochemistry were then used to verify the expression of the target proteins. Finally, logistic regression was performed to select serum proteins for the diagnostic model. As a result, five specific differentially expressed proteins, TGFß RIII, LAG-3, carboxypeptidase A2, Decorin and ANGPTL3, were found to have the ability to distinguish GC. Logistic regression analysis showed that the combination of carboxypeptidase A2 and TGFß RIII had superior potential for diagnosing GC (area under the ROC curve [AUC] = 0.801). The results suggested that these five proteins alone and the combination of carboxypeptidase A2 and TGFß RIII may be used as serum markers for the diagnosis of GC.
Assuntos
Biomarcadores Tumorais , Neoplasias Gástricas , Humanos , Análise Serial de Proteínas , Neoplasias Gástricas/diagnóstico , Carboxipeptidases A , Detecção Precoce de Câncer , Curva ROC , Proteína 3 Semelhante a AngiopoietinaRESUMO
Background: Researches showed RNA methylation genes can affect the prognosis of tumors. Thus, the study aimed to comprehensively analyze the effects of RNA methylation regulatory genes in prognosis and treatment of colorectal cancer (CRC). Methods: Prognostic signature associated with CRCs were constructed by differential expression analysis, Cox and Least Absolute Shrinkage and Selection Operator (LASSO) analyses. Receiver operating characteristic (ROC) and Kaplan-Meier survival analyses were used to validate the reliability of the developed model. Gene Ontology (GO), Gene set variation analysis (GSVA), and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed for functional annotation. Finally, normal and cancerous tissue were collected to validate gene by quantitative real-time PCR (qRT-PCR). Results: A prognostic risk model based on leucine rich pentatricopeptide repeat containing (LRPPRC) and ubiquitin-like with PHD and ring finger domains 2 (UHRF2) was constructed and relevant to the overall survival (OS) of CRC. Functional enrichment analysis revealed that collagen fibrous tissue, ion channel complex and other pathways were significantly enriched, which might help explain the underlying molecular mechanisms. There were significant differences in ImmuneScore, StromalScore, ESTIMATEScore between high- and low-risk groups (p < 0.05). Ultimately, qRT-PCR validation showed that a significant upregulation in the expression of LRPPRC and UHRF2 in cancerous tissue, which verified the effectiveness of our signature. Conclusion: In conclusion, 2 prognostic genes (LRPPRC and UHRF2) related to RNA methylation were identified by bioinformatics analysis, which might supply a new insight into the treatment and evaluation of CRC.
RESUMO
Esophageal squamous cell carcinoma (ESCC) is one of the most frequently occurring diseases in the world. Rabdosia rubescens (RR) has been demonstrated to be effective against ESCC; however, the mechanism is unknown. The primary gene modules related to the clinical characteristics of ESCC were initially investigated in this research using weighted gene co-expression network analysis (WCGNA) and differential expression gene (DEG) analysis. We employed network pharmacology to study the hub genes linked with RR therapy on ESCC. A molecular docking simulation was achieved to identify the binding activity of central genes to RR compounds. Lastly, a chain of experimentations was used to verify the inhibitory effect of RR water extract on the ESCC cell line in vitro. The outcomes revealed that CCNA2, TOP2A, AURKA, CCNB2, CDK2, CHEK1, and other potential central targets were therapeutic targets for RR treatment of ESCC. In addition, these targets are over-represented in several cancer-related pathways, including the cell cycle signaling pathway and the p53 signaling pathway. The predicted targets displayed good bonding activity with the RR bioactive chemical according to a molecular docking simulation. In vitro experiments revealed that RR water extracts could inhibit ESCC cells, induce cell cycle arrest, inhibit cell proliferation, increase P53 expression, and decrease CCNA2, TOP2A, AURKA, CCNB2, CDK2, and CHEK1. In conclusion, our study reveals the molecular mechanism of RR therapy for ESCC, providing great potential for identifying effective compounds and biomarkers for ESCC therapy.
RESUMO
BACKGROUND/AIMS: Epithelial cells line the intestinal mucosa and form an important barrier for maintaining host health. This study aimed to explore the mechanism of the Sphingosine-1-phosphate (S1P)/Sphingosine-1-phosphate receptor 2 (S1PR2) pathway in intestinal epithelial cells (IECs) that participate in the intestinal barrier function. METHODS: In this study, we constructed a knockout of the S1PR2 gene in mice, and Dextra sulfate sodium (DSS) was used to induce colitis. We isolated IECs from wild type (WT) and S1PR2-/- mice, and the endogenous expression of S1PR2 and Zonula occludens 1 (ZO-1) in IEC were detected by Western blot. Next, the major histocompatibility complex II (MHC-II) expression was analyzed by reverse transcription quantitative real-time (RT-qPCR) and flow cytometry. The in vivo and in vitro intestinal permeability were evaluated by serum fluorescein isothiocyanate (FITC) concentration. The tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interferon-γ (IFN-γ) levels in cell suspension were analyzed by enzyme-linked immuno sorbent assay (ELISA). A carboxyfluorescein diacetate succinimidyl ester (CFSE) assay was used to detect the T-cell proliferation in a co-culture system. RESULTS: The intestinal mucosal barrier damage in S1PR2-/- mice was more severe than in the WT mice, and there were more CD4+T-cells in the colon tissue of DSS-treated S1PR2-/- mice. Either the mouse colon carcinoma cell line (CT26. WT) or the IECs upregulated MHC-II expression, which then promoted CD4+T-cell proliferation. The S1P/S1PR2 pathway controlled MHC-II expression to regulate CD4+T-cell proliferation via the extracellular signal-regulated kinase (ERK) pathway. In addition, the IFN-γ that was secreted by CD4+T-cells increased DSS-induced damage of intestinal epithelial cell barrier function. ZO-1 expression was increased by S1P in CT26.WT cells, while S1PR2 antagonist JTE-013 expression was downregulated. However, in CT26.WTsi-S1PR2 cells, S1P had no effect on ZO-1 expression. CONCLUSIONS: The S1P/S1PR2 axis in IECs mediated CD4+T-cell activation via the ERK pathway and MHC-II expression to regulate intestinal barrier function.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Colite/imunologia , Mucosa Intestinal/imunologia , Lisofosfolipídeos/imunologia , Receptores de Lisoesfingolipídeo/imunologia , Transdução de Sinais , Esfingosina/análogos & derivados , Animais , Linfócitos T CD4-Positivos/patologia , Comunicação Celular , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Proliferação de Células , Células Cultivadas , Colite/genética , Colite/patologia , Células Epiteliais/imunologia , Células Epiteliais/patologia , Feminino , Absorção Intestinal , Mucosa Intestinal/patologia , Sistema de Sinalização das MAP Quinases , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Lisoesfingolipídeo/genética , Esfingosina/imunologia , Receptores de Esfingosina-1-FosfatoRESUMO
BACKGROUND: Aberrant expression of miRNAs was a critical element in the pathogenesis of inflammatory bowel disease (IBD). This study aimed to explore the involvement and mechanism of miR-126 in IBD. METHODS: In this study, the endogenous expressions of miR-126, S1PR2 and S1P in the pathological tissues of patients with IBD were detected using qRT-PCR and western blot assay, respectively. The luciferase reporter gene assay was performed to confirm the targeting regulatory relation between miR-126 and S1PR2. The transendothelial electrical resistance assay was used to measured the value of TEER. RESULTS: The expressions of miR-126, S1PR2 and S1P in the pathological tissues of IBD patients were significantly higher than that of the control group. Moreover, miR-126 overexpression contributed to intestinal mucosal barrier dysfunction in vitro. S1PR2 was a direct target of miR-126, and S1PR2 expression was negatively regulated by miR-126 in Caco-2 cells. However, S1PR2 activated by S1P had the protection effect for the integrity and permeability of intestinal mucosal barrier via a PI3K/Akt dependent mechanism. MiR-126 silencing possessed obvious protective effects on the intestinal barrier function, but these effects could be reversed by JTE-013 or LY294002. CONCLUSION: MiR-126 down-regulated S1PR2 and then prevented the activation of PI3K/AKT signaling pathway, which ultimately could damage intestinal mucosal barrier function.
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Absorção Intestinal/fisiologia , Mucosa Intestinal/fisiologia , MicroRNAs/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Lisoesfingolipídeo/metabolismo , Células CACO-2 , Humanos , Permeabilidade , Transdução de Sinais/fisiologia , Receptores de Esfingosina-1-FosfatoRESUMO
Sphingosine-1 phosphate (S1P) is a potent bioactive lipid mediator that acts both as an intracellular signaling molecule and a natural ligand of five different G protein-coupled receptors (GPCRs), S1PR1-5. The level of S1P in intestinal tissue is abundant. Previous studies have reported that S1P protects intestinal epithelial cell from apoptosis by activating the ERK and Akt signaling pathways. However, the effect of S1P on intestinal epithelial cell proliferation under physiological conditions and the underlying signaling mechanisms remain to be elucidated. Here, we show that, except for S1PR4, all S1PRs are expressed in normal intestinal epithelial cells with S1PR2 being the most abundant. S1P dose-dependently stimulated cell migration and proliferation, which were inhibited by JTE-013, a selective chemical antagonist of S1PR2, and by a S1PR2 shRNA. S1P significantly upregulated the expression of c-Myc, cyclin D1, E-cadherin and zona occluden-1 (ZO-1), which was completely inhibited by downregulation of S1PR2 expression with a shRNA. In total, the results suggest that S1P-mediated activation of the S1PR2 plays an important role in regulating intestinal epithelial cell proliferation and migration.
Assuntos
Mucosa Intestinal/citologia , Mucosa Intestinal/metabolismo , Lisofosfolipídeos/metabolismo , Receptores de Lisoesfingolipídeo/metabolismo , Esfingosina/análogos & derivados , Animais , Caderinas/genética , Linhagem Celular , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Ciclina D1/genética , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Regulação da Expressão Gênica , Genes myc , Sistema de Sinalização das MAP Quinases , Interferência de RNA , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/genética , Ratos , Receptores de Lisoesfingolipídeo/antagonistas & inibidores , Receptores de Lisoesfingolipídeo/genética , Esfingosina/metabolismo , Receptores de Esfingosina-1-Fosfato , Proteína da Zônula de Oclusão-1/genéticaRESUMO
Lymphangioma is an uncommon benign tumor that develops in the lymphatic system. Abdominal lymphangiomatosis is extremely rare in adult patients, and the clinical symptoms of this condition are complicated and atypical. We report a case of abdominal lymphangiomatosis in a 38-year-old female who presented with intestinal bleeding and protein-losing enteropathy, as well as lesions in the lung and bones. A computed tomography scan revealed multiple small cystic lesions without enhancement. Histological examination revealed microscopic cysts were submucosal, with walls composed of thin fibrous tissue, and D2-40 stained highlight the lining of the lymphatic channels by immunohistochemical method. We make a comparison with the cases reported before, and also discuss the diagnose of diffuse pulmonary lymphangiomatosis and Gorham's disease.
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Neoplasias Abdominais/diagnóstico , Linfangioma/diagnóstico , Neoplasias Abdominais/complicações , Neoplasias Abdominais/patologia , Adulto , Biópsia , Endoscopia Gastrointestinal , Feminino , Hemorragia Gastrointestinal/diagnóstico , Hemorragia Gastrointestinal/etiologia , Humanos , Imuno-Histoquímica , Pneumopatias/congênito , Pneumopatias/diagnóstico , Pneumopatias/etiologia , Linfangiectasia/congênito , Linfangiectasia/diagnóstico , Linfangiectasia/etiologia , Linfangioma/etiologia , Linfangioma/patologia , Osteólise Essencial/diagnóstico , Osteólise Essencial/etiologia , Enteropatias Perdedoras de Proteínas/diagnóstico , Enteropatias Perdedoras de Proteínas/etiologia , Tomografia Computadorizada por Raios XRESUMO
BACKGROUND/AIMS: H. pylori persists for the virtual life of its host. Recent studies suggested that CD4 CD25+ regulatory T cells may be involved in this process. However, the alteration of CD4+ CD25+ regulatory T cells after eradication of H. pylori remains a question. METHODOLOGY: By using biopsies from 45 H. pylori-positive patients and the ones after eradication of H. pylori and 35 H. pylori-negative adults, real-time PCR and general PCR were used to quantify the expression of Foxp3 mRNA. IHC was used to semi- quantify the number of CD4+ CD25+ T cells in gastric mucosa. RESULTS: We found that proportion ofCD25+ T cell in CD4+ T cells accounted for 0.739% in H. pylori-negative individuals, while it was accounted for 5.012% in H. pylori-positive patients. After eradication of H. pylori, proportion of CD25+ T cell in CD4+ T cells declined (P mRNA significantly decreased (P < 0.01) in gastric mucosa of patients after eradication of H. pylori. CONCLUSIONS: CD4+ CD25+ regulatory T cells decreased in gastric mucosa when patients received eradication of H. pylori. Eradication of H. pylori results in the significant decrease of Foxp3 mRNA in gastric mucosal, or using the drugs of anti-H. pylori induce the reduction of gastric mucosal Foxp3 mRNA expression, which is the a key regulatory gene for the development and function of CD4+ CD25+ regulatory T cells, thus contributing to the eradication of H. pylori. All the data offer new possibilities that Foxp3 gene may be the new target of immunization intervention strategies for eradication of H. pylori.
Assuntos
Antígenos CD4/biossíntese , Fatores de Transcrição Forkhead/biossíntese , Infecções por Helicobacter/metabolismo , Helicobacter pylori/isolamento & purificação , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Linfócitos T Reguladores/metabolismo , Adulto , Biópsia , Estudos de Coortes , Feminino , Fatores de Transcrição Forkhead/genética , Mucosa Gástrica/química , Mucosa Gástrica/citologia , Infecções por Helicobacter/imunologia , Infecções por Helicobacter/patologia , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/análise , RNA Mensageiro/genética , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/imunologiaRESUMO
BACKGROUND: Fibrinogen-like protein 2 (FGL2), a new member of the fibrinogen-like family, has recently been identified as a novel immunosuppressive molecule. AIM: The purpose of this work was to investigate intestinal and peripheral expression of FGL2 in patients with inflammatory bowel disease (IBD), mainly ulcerative colitis (UC) and Crohn's disease (CD). METHODS: FGL2 expression in mucosal biopsies from three groups (UC group (n = 61), CD group (n = 54), and controls group (n = 35)) was detected by immunohistochemistry. Concentrations of FGL2 in plasma from 50 UC patients, 45 CD patients, and 30 controls were analyzed by enzyme-linked immunosorbent assay. Western blot of FGL2 protein and real-time fluorescent quantitative PCR of FGL2 mRNA expression by peripheral mononuclear cells was performed. Correlations of FGL2 expression with disease type, activity, and location, and with measured laboratory data, including C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR), were examined. RESULTS: Intestinal and peripheral FGL2 protein data showed that FGL2 expression was significantly up-regulated in both UC and CD patients compared with controls (P < 0.001). Expression of FGL2 was higher in UC and CD patients with active disease than in those with inactive disease (P < 0.001). Moreover, FGL2 mRNA expression was significantly higher in patients with active disease than in those with inactive disease (P < 0.050). Expression of FGL2 protein was correlated with disease activity indices, CRP levels, and ESR levels. CONCLUSION: Expression of FGL2 was up-regulated in IBD patients with active disease. Measurement of FGL2 may be used as a helpful biomarker for understanding immunopathogenesis and for assessment of IBD.