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The heterogeneity of hepatocellular carcinoma (HCC) can prevent effective treatment, emphasizing the need for more effective therapies. Herein, we employed arsenene nanosheets coated with manganese dioxide and polyethylene glycol (AMPNs) for the degradation of Pin1, which is universally overexpressed in HCC. By employing an "AND gate", AMPNs exhibited responsiveness toward excessive glutathione and hydrogen peroxide within the tumor microenvironment, thereby selectively releasing AsxOy to mitigate potential side effects of As2O3. Notably, AMPNs induced the suppressing Pin1 expression while simultaneously upregulation PD-L1, thereby eliciting a robust antitumor immune response and enhancing the efficacy of anti-PD-1/anti-PD-L1 therapy. The combination of AMPNs and anti-PD-1 synergistically enhanced tumor suppression and effectively induced long-lasting immune memory. This approach did not reveal As2O3-associated toxicity, indicating that arsenene-based nanotherapeutic could be employed to amplify the response rate of anti-PD-1/anti-PD-L1 therapy to improve the clinical outcomes of HCC patients and potentially other solid tumors (e.g., breast cancer) that are refractory to anti-PD-1/anti-PD-L1 therapy.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Compostos de Manganês , Peptidilprolil Isomerase de Interação com NIMA , Óxidos , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologia , Óxidos/química , Óxidos/farmacologia , Humanos , Peptidilprolil Isomerase de Interação com NIMA/antagonistas & inibidores , Peptidilprolil Isomerase de Interação com NIMA/metabolismo , Compostos de Manganês/química , Compostos de Manganês/farmacologia , Nanoestruturas/química , Antineoplásicos/química , Antineoplásicos/farmacologia , Arsenicais/química , Arsenicais/farmacologia , Arsenicais/uso terapêutico , Camundongos , Animais , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Linhagem Celular Tumoral , Polietilenoglicóis/químicaRESUMO
The NLRP3 inflammasome functions as an inflammatory driver, but its relationship with lipid metabolic changes in early sepsis remains unclear. Here, we found that GITR expression in monocytes/macrophages was induced by lysophosphatidylcholine (LPC) and was positively correlated with the severity of sepsis. GITR is a costimulatory molecule that is mainly expressed on T cells, but its function in macrophages is largely unknown. Our in vitro data showed that GITR enhanced LPC uptake by macrophages and specifically enhanced NLRP3 inflammasome-mediated macrophage pyroptosis. Furthermore, in vivo studies using either cecal ligation and puncture (CLP) or LPS-induced sepsis models demonstrated that LPC exacerbated sepsis severity/lethality, while conditional knockout of GITR in myeloid cells or NLRP3/caspase-1/IL-1ß deficiency attenuated sepsis severity/lethality. Mechanistically, GITR specifically enhanced inflammasome activation by regulating the posttranslational modification (PTM) of NLRP3. GITR competes with NLRP3 for binding to the E3 ligase MARCH7 and recruits MARCH7 to induce deacetylase SIRT2 degradation, leading to decreasing ubiquitination but increasing acetylation of NLRP3. Overall, these findings revealed a novel role of macrophage-derived GITR in regulating the PTM of NLRP3 and systemic inflammatory injury, suggesting that GITR may be a potential therapeutic target for sepsis and other inflammatory diseases. GITR exacerbates LPC-induced macrophage pyroptosis in sepsis via posttranslational regulation of NLRP3. According to the model, LPC levels increase during the early stage of sepsis, inducing GITR expression on macrophages. GITR not only competes with NLRP3 for binding to the E3 ligase MARCH7 but also recruits MARCH7 to induce the degradation of the deacetylase SIRT2, leading to decreasing ubiquitination but increasing acetylation of NLRP3 and therefore exacerbating LPC-induced NLRP3 inflammasome activation, macrophage pyroptosis and systemic inflammatory injury.
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Proteína Relacionada a TNFR Induzida por Glucocorticoide , Lisofosfatidilcolinas , Macrófagos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR , Processamento de Proteína Pós-Traducional , Piroptose , Sepse , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Animais , Sepse/imunologia , Macrófagos/metabolismo , Macrófagos/imunologia , Lisofosfatidilcolinas/metabolismo , Camundongos , Proteína Relacionada a TNFR Induzida por Glucocorticoide/metabolismo , Inflamassomos/metabolismo , Masculino , Camundongos Knockout , Humanos , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Sirtuína 2/metabolismo , Sirtuína 2/genética , AcetilaçãoRESUMO
BACKGROUND: Many studies have found that glucocorticoid (GC) combined with hydroxychloroquine (HCQ) has a good clinical effect in the treatment of systemic lupus erythematosus (SLE) rash, but there is no relevant systematic evaluation at present. The purpose of this study was to systematically evaluate and analyze the effectiveness and safety of GC combined with HCQ in the treatment of SLE rash. METHODS: Randomized controlled trials of GC combined with HCQ in the treatment of SLE rash were collected through computer retrieval of Cochrane Library, PubMed, Embase, CNKI, China Science and Technology Journal Database (VIP), Wanfang Data Knowledge Service Platform (Wanfang), and China Biology Medicine disc (CBM) since the establishment of the database. The main outcome indicators included clinical total effective rate, adverse reactions, SLE disease activity index (SLEDAI) score, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and complement 3 (C3). A meta-analysis was conducted using Review Manager 5.3 software. RESULTS: A total of 11 studies involving 809 patients (406 in the test group and 403 in the control group) were included in this article. The meta-analysis results showed that compared with the single use of GC, GC combined with HCQ could improve the clinical total effective rate in the treatment of SLE rash (odds ratio [OR] = 4.27, 95% confidence interval [CI] 2.50-7.30, p < .00001), and reduce the occurrence of adverse reactions (OR = 0.26, 95% CI 0.15-0.44, p < .00001); effectively reduce SLEDAI score (mean difference [MD] = 1.88, 95% CI 1.66-2.10, p < .00001) and ESR level (MD = 7.92, 95% CI 5.66-10.19, p < .00001); increase C3 level after treatment (MD = 0.36, 95% CI 0.32-0.41, p < .00001); and reduce CRP level (MD = 3.22, 95% CI 2.87-3.58, p < .00001), with statistically significant differences. CONCLUSION: Compared with the use of GC alone, GC combined with HCQ can improve the clinical effectiveness of SLE rash treatment, with a low incidence of adverse reactions and good clinical safety. However, the number and quality of studies included in this article were not high, so the findings need to be further verified by high-quality, multicenter randomized controlled trials.
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Exantema , Lúpus Eritematoso Sistêmico , Humanos , Hidroxicloroquina/efeitos adversos , Glucocorticoides/efeitos adversos , Lúpus Eritematoso Sistêmico/diagnóstico , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Lúpus Eritematoso Sistêmico/induzido quimicamente , Resultado do Tratamento , Exantema/induzido quimicamente , Exantema/diagnóstico , Exantema/tratamento farmacológico , Estudos Multicêntricos como AssuntoRESUMO
Metabolic dysregulation contributes not only to cancer development but also to a tumor immune microenvironment (TIME), which poses great challenges to chemo- and immunotherapy. Targeting metabolic reprogramming has recently emerged as a promising strategy for cancer treatment, but the lethality against solid tumors appears to be fairly restricted, partially due to the poor solubility of small molecule drugs. Herein, we construct a versatile biomimetic nanoplatform (referred to as HM-BPT) employing pH-sensitive tumor-tropism hybrid membrane-coated Manganese oxide (MnO2) nanoparticles for the delivery of BPTES, a glutamine metabolism inhibitor. Basically, hybrid membranes consisting of mesenchymal stem cell membranes (MSCm) and pH-sensitive liposomes (pSL) enable the biomimetic nanoplatform to target TME and escape from endo/lysosomes after endocytosis. The results reveal that HM-BPT treatment leads to remarkable tumor inhibition, cytotoxic T lymphocyte (CTL) infiltration, as well as M1 phenotype repolarization and stimulator of IFN genes (STING) pathway activation in macrophages in a 4T1 xenograft model. Furthermore, glutathione (GSH) depletion and oxygen (O2) supply synergistically ameliorate the immunosuppressive status of the TME, boosting potent antitumor immune responses. Overall, our study explores an integrated therapeutic platform for TME reprogramming and immune activation, offering tremendous promise for cancer combination therapy. STATEMENT OF SIGNIFICANCE: Metabolic abnormalities and the tumor immune microenvironment (TIME) lead to hyporesponsiveness to conventional therapies, ultimately resulting in refractory malignancies. In the current work, a biomimetic nanoplatform (HM-BPT) was developed for TME metabolic reprogramming in favor of immunotherapy. Particularly, hybrid membrane camouflage endowed the nanoplatform with TME targeting, endo/lysosomal escape, and sensitive release properties. The impact of hybrid membrane fusion ratio on cellular uptake and cell viability was explored, yielding beneficial references for the future development of bioactive nanomaterials. Intravenous administration of HM-BPT substantially relieved tumor burden and restored innate and acquired immune activation in 4T1 xenograft models. In conclusion, the created HM-BPT system has the potential to be a promising nanoplatform for combining cancer therapies.
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Nanopartículas , Neoplasias , Humanos , Animais , Compostos de Manganês/farmacologia , Microambiente Tumoral , Óxidos , Lisossomos , Imunoterapia , Modelos Animais de Doenças , Concentração de Íons de Hidrogênio , Linhagem Celular TumoralRESUMO
INTRODUCTION: Systemic lupus erythematosus (SLE) is a chronic autoimmune disease. Lupus nephritis (LN) is a common type of organ damage which occurs in SLE patients and is characterized by recurrent proteinuria. Activation of B lymphocytes can lead to refractory LN, which is an important pathogenic factor in SLE. B lymphocyte stimulator (BLyS) and A proliferation-inducing ligand (APRIL) are predominantly produced by myeloid cells (monocytes, dendritic cells, neutrophils, etc) to regulate B lymphocyte function. Telitacicept was the first dual-targeting biological drug which targeted both BLyS and APRIL. Telitacicept has passed a phase II clinical trial and has since been approved for the treatment of SLE. CASE PRESENTATION: We report a case of SLE confirmed by renal biopsy as proliferative lupus nephritis (PLN) with massive proteinuria, which was treated with telitacicept (European League Against Rheumatism / American College of Rheumatology 2019 standard). During the 19 months of follow-up, the patient's renal function was stable, massive proteinuria was relieved, and creatinine and blood pressure did not increase. CONCLUSIONS: During the 19 months of telitacicept treatment (160 mg once weekly), PLN reduced blood system damage and proteinuria without increasing the risk of infection.
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Lúpus Eritematoso Sistêmico , Nefrite Lúpica , Humanos , Nefrite Lúpica/diagnóstico , Nefrite Lúpica/tratamento farmacológico , Nefrite Lúpica/patologia , Proteinúria/tratamento farmacológico , Proteinúria/etiologia , BrancosRESUMO
PURPOSE: The purpose of this study was to assess the role of leukocyte immunoglobulin-like receptor A5 (LILRA5) in regulating bacterial infection and corneal inflammation. METHODS: The human corneal tissue microarray data set GSE58291 from Gene Expression Omnibus was downloaded. Then, the differentially expressed genes, Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, Gene Set Enrichment Analysis, and the immune infiltration analysis were conducted. We constructed the Pseudomonas aeruginosa ( P. aeruginosa ) keratitis mice model using wild-type and LILRA5-deficient mice. The results of the bioinformatics analysis were verified by the cell in vitro and animal in vivo experiments. RESULTS: This study revealed that LILRA5 is substantially expressed in human keratitis and regulates the immune response negatively. Neutrophils were identified as the core fraction of immune cells in keratitis. After P. aeruginosa infection, neutrophils lacking LILRA5 induced elevated levels of proinflammatory cytokines and toll-like receptor 4. LILRA5 deficiency exacerbated the severity of the infection and the production of proinflammatory cytokines in mice. CONCLUSIONS: LILRA5 was discovered as an immunosuppressive regulator in P. aeruginosa keratitis, highlighting its significance in activated immune responses.
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Ceratite , Infecções por Pseudomonas , Receptores Imunológicos , Animais , Humanos , Camundongos , Córnea/patologia , Citocinas/metabolismo , Imunoglobulinas/metabolismo , Ceratite/microbiologia , Camundongos Endogâmicos C57BL , Neutrófilos , Pseudomonas aeruginosa , Infecções por Pseudomonas/microbiologia , Receptores Imunológicos/genéticaRESUMO
Tumor-derived extracellular vesicle (tEV) biomarkers can reflect cancer cell phenotypes and have great potential for cancer diagnosis and treatment. However, tEVs display high heterogeneity, and rapid and sensitive identification of EV biomarkers remains challenging due to their low expression. Spectral overlap also significantly limits the multiplex analysis of EV biomarkers by fluorescent probes. Herein, we developed a method for highly sensitive tEV phenotyping that uses size-coded microbeads that carry hairpin probes that can bind to aptamers targeting distinct tEV biomarkers. We also designed a microfluidic chip containing spacer arrays that segregate these microbeads in distinct chip regions according to their size to generate location-specific signals indicating the level of different EV biomarkers. The EV biomarker signal on these microbeads was amplified by in situ rolling cyclic amplification (RCA). This strategy permits the simultaneous detection of multiple tEV phenotypes by fluorescence spectroscopy without the limitations of spectral overlap. This study demonstrates that this tEV phenotyping method can rapidly and simultaneously detect six different tEV phenotypes with high sensitivity. Due to the programmability of the sensing platform, this method can be rapidly adapted to detect different tEV phenotype substitutions of the detected biomarkers. Notably, clinical cohort studies show that this strategy may provide new ideas for the precise diagnosis and personalized treatment of cancer patients.
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Vesículas Extracelulares , Neoplasias , Humanos , Microesferas , Fenótipo , Biomarcadores Tumorais/metabolismo , Neoplasias/metabolismo , Vesículas Extracelulares/químicaRESUMO
Triggering receptors expressed on myeloid cells 2 (TREM2) is considered a protective factor to protect host from bacterial infection, while how it elicits this role is unclear. In the present study, we demonstrate that deficiency of triggering receptors expressed on myeloid cells 2 (TREM2) significantly enhanced macrophage pyroptosis induced by four common pyogenic bacteria including Staphylococcus aureus, Pseudomonas aeruginosa, Streptococcus pneumoniae, and Escherichia coli. TREM2 deficiency also decreased bacterial killing ratio of macrophage, while Caspase-1 or GSDMD inhibition promoted macrophage-mediated clearance to these bacteria. Further study demonstrated that the effect of TREM2 on macrophage pyroptosis and bacterial eradication mainly dependents on the activated status of NLRP3 inflammasome. Moreover, as the key downstream of TREM2, ß-catenin phosphorylated at Ser675 by TREM2 signal and accumulated in nucleus and cytoplasm. ß-catenin mediated the effect of TREM2 on NLRP3 inflammasome and macrophage pyroptosis by reducing NLRP3 expression, and inhibiting inflammasome complex assembly by interacting with ASC. Collectively, TREM2/ß-catenin inhibits NLRP3 inflammasome to regulate macrophage pyroptosis, and enhances macrophage-mediated pyogenic bacterial clearance.
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Inflamassomos , Piroptose , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Inflamassomos/metabolismo , Macrófagos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Pseudomonas aeruginosa , beta Catenina/metabolismoRESUMO
Stimulator of interferon genes (STING) is a cytosolic DNA sensor which is regarded as a potential target for antitumor immunotherapy. However, clinical trials of STING agonists display limited anti-tumor effects and dose-dependent side-effects like inflammatory damage and cell toxicity. Here, we showed that tetrahedral DNA nanostructures (TDNs) actively enter macrophages to promote STING activation and M1 polarization in a size-dependent manner, and synergized with Mn2+ to enhance the expressions of IFN-ß and iNOS, as well as the co-stimulatory molecules for antigen presentation. Moreover, to reduce the cytotoxicity of Mn2+, we constructed a TDN-MnO2 complex and found that it displayed a much higher efficacy than TDN plus Mn2+ to initiate macrophage activation and anti-tumor response both in vitro and in vivo. Together, our studies explored a novel immune activation effect of TDN in cancer therapy and its synergistic therapeutic outcomes with MnO2. These findings provide new therapeutic opportunities for cancer therapy.
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Immune-checkpoint blockade is widely studied for cancer therapy. Although the co-inhibitory receptor Programmed death-1(PD-1) blockade benefits some non-small cell lung cancer (NSCLC) patients, a large portion of NSCLC patients still fail to respond to this immunotherapy, and the underlying mechanism is unclear. Thus, a synergistic therapy to enhance the effect of PD-1 is urgently needed to improve the poor outcome of NSCLC patients. Here, we demonstrated that effector memory T cells were increased and T cell response became stronger in PD-1 immunotherapy responders (n = 20) but not in non-responders (n = 10). The expression of co-stimulatory receptor OX40 was upregulated on T cells following PD-1 immunotherapy and was positively associated with the percentage of PD-1+T cells and the responsiveness of T cells. Combination treatment of antagonistic anti-PD-1 and agonistic anti-OX40 antibodies (Abs) promoted the proliferation and cytokines production of T cells from PBMCs of non-responders ex vivo. Consistently, anti-PD-1 and anti-OX40 therapy synergistically augmented T cell response in an in vivo mouse lung cancer model. Our study confirmed the antitumor effects of anti-PD-1/OX40 combination in lung cancer patients as well as in the murine lung cancer model, and the results provide a rationale for clinical trials evaluating the therapeutic effect of this combination of antibodies for NSCLC immunotherapy.
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Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Animais , Anticorpos/uso terapêutico , Antígeno B7-H1 , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Humanos , Imunidade , Imunoterapia/métodos , Neoplasias Pulmonares/tratamento farmacológico , Camundongos , Receptor de Morte Celular Programada 1RESUMO
The coronavirus disease 2019 (COVID-19) emerged around December 2019 and have become a global epidemic disease currently. Specific antibodies against SAS-COV-2 could be detected in COVID-19 patients' serum or plasma, but the clinical values of these antibodies as well as the effects of clinical drugs on humoral responses have not been fully demonstrated. In this study, 112 plasma samples were collected from 36 patients diagnosed with laboratory-confirmed COVID-19 in the Fifth Affiliated Hospital of Sun Yat-sen University. The IgG and IgM antibodies against receptor binding domain (RBD) and spike protein subunit 1 (S1) of SAS-COV-2 were detected by ELISA. We found that COVID-19 patients generated specific antibodies against SARS-CoV-2 after infection, and the levels of anti-RBD IgG within 2 to 3 weeks from onset were negatively associated with the time of positive-to-negative conversion of SARS-CoV-2 nucleic acid. Patients with severe symptoms had higher levels of anti-RBD IgG in 2 to 3 weeks from onset. The use of chloroquine did not significantly influence the patients' antibody titer but reduced C-reaction protein (CRP) level. Using anti-viral drugs (lopinavir/ritonavir or arbidol) reduced antibody titer and peripheral lymphocyte count. While glucocorticoid therapy developed lower levels of peripheral lymphocyte count and higher levels of CRP, lactate dehydrogenase (LDH), α-Hydroxybutyrate dehydrogenase(α-HBDH), total bilirubin (TBIL), direct bilirubin (DBIL). From these results, we suggested that the anti-RBD IgG may provide an early protection of host humoral responses against SAS-COV-2 infection within 2 to 3 weeks from onset, and clinical treatment with different drugs displayed distinct roles in humoral and inflammatory responses.
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COVID-19/imunologia , Indóis/uso terapêutico , Lopinavir/uso terapêutico , Ritonavir/uso terapêutico , SARS-CoV-2/fisiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/sangue , Formação de Anticorpos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Glicoproteína da Espícula de Coronavírus/imunologia , Tratamento Farmacológico da COVID-19RESUMO
Circulating tumor cell (CTC) clusters, which are multicellular groups of CTCs, were recently suggested to had the greater potential of forming distal metastasis than single CTCs. However, our understanding of the forming of CTC clusters is still limited since there are few existing methods to study cancer cells aggregation kinetics, especially for a small number of cells. Herein we report a high-throughput miniaturized microwell-based cell aggregation-chip (AG-chip) to enable better characterize of the tumor cells clustering process. We successfully demonstrated the capability of the AG-chip in determining cell aggregation, and found that: (1) high metastatic breast cancer cells (MDA-MB-231 & MDA-MB-436) have stronger aggregation capacities than those low metastatic breast cancer cells (MCF-7 & SK-BR-3); (2) cells with similar aggregation ability were distinguished through the analysis of aggregation kinetics; (3) the detected aggregation ability can be used to indicate the metastatic potential of the cells; (4) the inhibition of integrins could regulate the cell clustering via blockage of cell adhesion or/and cell migration. This newly developed microdevice may promote further study of CTC clusters and metastasis.
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Neoplasias da Mama , Células Neoplásicas Circulantes , Contagem de Células , Linhagem Celular Tumoral , Análise por Conglomerados , Humanos , Metástase Neoplásica , Células Neoplásicas Circulantes/patologiaRESUMO
Pore-forming proteins are an agent for attack or defense in various organisms, and its cytolytic activity has medical potential in cancer therapy. Despite recent advances in mimicking these proteins by amphipathic DNA nanopores, it remains inefficient to incorporate them into lipid bilayers. Here, we present the development of vesicular DNA nanopores that can controllably open a lipid membrane. Different from previously reported DNA nanopores that randomly insert into the planar bilayers, we design on-command fusogenic liposome-incorporated transmembrane DNA nanopores (FLIPs) that bypass the direct insertion process. By steric deshielding of fusogenic liposomal supports under low pH conditions, the embedded FLIPs are transferred and perforate lipid bilayers. We find that FLIPs depolarize the plasma membrane and thereby induce pyroptosis-like cell death. We further demonstrate the use of FLIPs to inhibit tumor growth in murine tumor models, which provides a new route to cancer nanotherapy.
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Membrana Celular/metabolismo , DNA , Bicamadas Lipídicas , Nanoporos , Piroptose/efeitos dos fármacos , Células A549 , Animais , Membrana Celular/patologia , DNA/química , DNA/farmacologia , Humanos , Bicamadas Lipídicas/química , Bicamadas Lipídicas/farmacologia , Lipossomos , CamundongosRESUMO
Mucosal-associated invariant T (MAIT) cells play a key role in local and systemic immune responses. Studies suggest that type 2 diabetes (T2D) is associated with alterations in the human MAIT cell response. However, the mechanisms that regulate the survival and homeostasis of human MAIT cells are poorly defined. In this study, we demonstrate that the costimulatory TNF superfamily receptor OX40 was highly expressed in MAIT cells of patients with T2D. Compared with OX40-negative MAIT cells, OX40-positive MAIT cells showed a high activation and a memory phenotype. Surprisingly, OX40 expression was negatively correlated with the frequency of MAIT cells in the peripheral blood of T2D patients. Increased cleaved caspase-3 levels were observed in OX40+-expressing MAIT cells in T2D patients. In vitro, activated OX40 signaling by recombinant OX40L protein promoted caspase-3 activation and apoptosis of MAIT cells. Inhibition of caspase-3 restored apoptosis of MAIT cells induced by OX40 signaling. These results identify OX40 as an amplifier of activation-induced cell death of human blood MAIT cells and shed new light on the regulation of MAIT cells in the phase of immune responses in T2D.
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Diabetes Mellitus Tipo 2/sangue , Células T Invariantes Associadas à Mucosa/metabolismo , Receptores OX40/metabolismo , Adulto , Apoptose/efeitos dos fármacos , Caspase 3/metabolismo , Estudos de Coortes , Feminino , Humanos , Memória Imunológica , Ativação Linfocitária/imunologia , Masculino , Pessoa de Meia-Idade , Células T Invariantes Associadas à Mucosa/imunologia , Ligante OX40/farmacologia , Fenótipo , Proteínas Recombinantes/farmacologia , Transdução de Sinais/efeitos dos fármacosRESUMO
Efforts have been taken to enhance the study of single-cells, however, the task remains challenging because most previous investigations cannot exclude the interactions between single cells or separately retrieved cells with specificity for further analyses. Here, a single-cell mobility analysis platform (SCM-Chip) is developed that can not only real-time monitor single-cell migration in independent niches but can also selectively recover target cells one by one. The design of each channel with a single-cell capture unit and an outlet enables the system to place single cells in different isolated niches with fluidic capture and to respectively collect target cells based on mobilities. SCM-Chip characterization of breast cancer cells reveals the presence of high- and low-migratory populations. Whole-cell transcriptome analysis establishes that monocyte chemotactic protein induced protein 1 (MCPIP1) is related with cell mobility; cells with a high expression of MCPIP1 exhibit low mobility in vitro and metastasis in vivo. The SCM platform provides a generic tool for accurate single-cell isolation and differentiation that can be readily adapted for the study of cancer and drug development.
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Pseudomonas aeruginosa (PA) is the leading cause of bacterial keratitis, especially in those who wear contact lens and who are immunocompromised. Once the invading pathogens are recognized by pattern recognition receptors expressed on the innate immune cells, the innate immune response is stimulated to exert host defense function, which is the first line to fight against PA infection. As a converging point of cytosolic DNA sense signaling, stimulator of interferon genes (STING) was reported to participate in host-pathogen interaction. However, the role of STING in regulating PA-induced corneal inflammation and bacterial clearance remains unknown. Our data demonstrated that STING was activated in murine model of PA keratitis and in in vitro-cultured macrophages, indicated by Western blot, immunostaining, and flow cytometry. To explore the role of STING in PA keratitis, we used siRNA to silence STING and 2',3'-cGAMP to activate STING in vivo and in vitro, and the in vivo data found out that STING promoted host resistance against PA infection. To investigate the reason why STING played a protective role in PA keratitis, the inflammatory cytokine secretion and bacterial load were measured by using real-time PCR and bacterial plate count, respectively. Our data demonstrated that STING suppressed the production of inflammatory cytokines and enhanced bacterial elimination in murine model of PA keratitis and in PA-infected macrophages. To further investigate the mechanism beneath, the phosphorylation of mitogen-activated protein kinase, the nuclear translocation of nuclear factor-κB (NF-κB) and the bactericidal mechanism were measured by western-blot, immunofluorescence, and real-time PCR, respectively. Our data indicated that STING suppressed inflammatory cytokine expressing via restraining NF-κB activity and enhanced inducible NO synthase expression, an oxygen-dependent bactericidal mechanism. In conclusion, this study demonstrated that STING promoted host resistance against PA keratitis and played a protective role in PA-infected corneal disease, via inhibiting corneal inflammation and enhancing bacterial killing.