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1.
Nat Commun ; 12(1): 5029, 2021 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-34413303

RESUMO

Dendritic cells (DC) in the lung that induce Th17 differentiation remain incompletely understood, in part because conventional CD11b+ DCs (cDC2) are heterogeneous. Here, we report a population of cDCs that rapidly accumulates in lungs of mice following house dust extract inhalation. These cells are Ly-6C+, are developmentally and phenotypically similar to cDC2, and strongly promote Th17 differentiation ex vivo. Single cell RNA-sequencing (scRNA-Seq) of lung cDC2 indicates 5 distinct clusters. Pseudotime analysis of scRNA-Seq data and adoptive transfer experiments with purified cDC2 subpopulations suggest stepwise developmental progression of immature Ly-6C+Ly-6A/E+ cDC2 to mature Ly-6C-CD301b+ lung resident cDC2 lacking Ccr7 expression, which then further mature into CD200+ migratory cDC2 expressing Ccr7. Partially mature Ly-6C+Ly-6A/E-CD301b- cDC2, which express Il1b, promote Th17 differentiation. By contrast, CD200+ mature cDC2 strongly induce Th2, but not Th17, differentiation. Thus, Th17 and Th2 differentiation are promoted by lung cDC2 at distinct stages of maturation.


Assuntos
Asma/imunologia , Antígeno CD11b/imunologia , Células Dendríticas/imunologia , Pulmão/imunologia , Células Th17/imunologia , Células Th2/imunologia , Transferência Adotiva/métodos , Animais , Asma/metabolismo , Asma/patologia , Sequência de Bases , Antígeno CD11b/metabolismo , Diferenciação Celular/fisiologia , Células Cultivadas , Técnicas de Cocultura , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Pulmão/metabolismo , Pulmão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Análise de Célula Única/métodos , Células Th17/citologia , Células Th2/citologia
2.
Oncoimmunology ; 10(1): 1940676, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34239774

RESUMO

The optimization of adoptive transfer approaches of anti-tumor T cells requires both the functional improvement of the injected T cells and the modulation of the tumor microenvironment, favoring the recruitment of these T cells and their activation. We have recently shown the therapeutic benefit of two approaches tested individually in a melanoma model wich were on one hand the adoptive transfer of specific T cells deficient for the expression of the inhibitory receptor PD-1, and on the other hand PD-L1 targeted alpha therapy (TAT). In this study, we sought to investigate the efficacy of these two therapies combined, compared to each monotherapy, in order to evaluate the synergy between these two approaches, in the same melanoma model. Here we used melanoma-specific T-cell clones, previously validated for the edition of PDCD1 gene and with previously demonstrated superior anti-tumor activity than their wild-type counterparts, after adoptive transfer in NSG mice engrafted with PD-L1 expressing human melanoma tumors. We also used a previously validated TAT approach, using a 213Bi-anti-human-PD-L1 mAb, alone or in combination with adoptive cell transfer, in the same mouse model. We confirmed previous results obtained with each monotherapy and documented the safety and the superior ability of a combination between the adoptive transfer of PD-1 deficient T cells and TAT targeting PD-L1 to control the growth of melanoma tumors in NSG mice. This study provides the first proof-of-concept of the efficacy of a combination therapy using TAT, adoptive cell transfer and genomic editing of IC-coding genes.


Assuntos
Antígeno B7-H1 , Melanoma , Transferência Adotiva , Animais , Antígeno B7-H1/genética , Humanos , Melanoma/genética , Camundongos , Receptor de Morte Celular Programada 1/genética , Linfócitos T , Microambiente Tumoral
4.
Nat Commun ; 12(1): 4474, 2021 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-34294714

RESUMO

Non-alcoholic steatohepatitis (NASH) is a leading cause of chronic liver disease that can progress to liver fibrosis. Recent clinical advance suggests a reversibility of liver fibrosis, but the cellular and molecular mechanisms underlying NASH resolution remain unclarified. Here, using a murine diet-induced NASH and the subsequent resolution model, we demonstrate direct roles of CD8+ tissue-resident memory CD8+ T (CD8+ Trm) cells in resolving liver fibrosis. Single-cell transcriptome analysis and FACS analysis revealed CD69+CD103-CD8+ Trm cell enrichment in NASH resolution livers. The reduction of liver CD8+ Trm cells, maintained by tissue IL-15, significantly delayed fibrosis resolution, while adoptive transfer of these cells protected mice from fibrosis progression. During resolution, CD8+ Trm cells attracted hepatic stellate cells (HSCs) in a CCR5-dependent manner, and predisposed activated HSCs to FasL-Fas-mediated apoptosis. Histological assessment of patients with NASH revealed CD69+CD8+ Trm abundance in fibrotic areas, further supporting their roles in humans. These results highlight the undefined role of liver CD8+ Trm in fibrosis resolution.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Células Estreladas do Fígado/patologia , Cirrose Hepática/imunologia , Cirrose Hepática/patologia , Hepatopatia Gordurosa não Alcoólica/imunologia , Hepatopatia Gordurosa não Alcoólica/patologia , Transferência Adotiva , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Apoptose/imunologia , Modelos Animais de Doenças , Progressão da Doença , Feminino , Células Estreladas do Fígado/imunologia , Humanos , Memória Imunológica , Interleucina-15/imunologia , Cirrose Hepática/terapia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Hepatopatia Gordurosa não Alcoólica/terapia , Receptores CCR5/imunologia , Adulto Jovem
5.
Nat Commun ; 12(1): 4365, 2021 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-34272369

RESUMO

Activating RAS missense mutations are among the most prevalent genomic alterations observed in human cancers and drive oncogenesis in the three most lethal tumor types. Emerging evidence suggests mutant KRAS (mKRAS) may be targeted immunologically, but mKRAS epitopes remain poorly defined. Here we employ a multi-omics approach to characterize HLA class I-restricted mKRAS epitopes. We provide proteomic evidence of mKRAS epitope processing and presentation by high prevalence HLA class I alleles. Select epitopes are immunogenic enabling mKRAS-specific TCRαß isolation. TCR transfer to primary CD8+ T cells confers cytotoxicity against mKRAS tumor cell lines independent of histologic origin, and the kinetics of lytic activity correlates with mKRAS peptide-HLA class I complex abundance. Adoptive transfer of mKRAS-TCR engineered CD8+ T cells leads to tumor eradication in a xenograft model of metastatic lung cancer. This study validates mKRAS peptides as bona fide epitopes facilitating the development of immune therapies targeting this oncoprotein.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Carcinogênese/imunologia , Epitopos de Linfócito T/imunologia , Neoplasias Pulmonares/imunologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Transferência Adotiva , Alelos , Animais , Carcinogênese/genética , Linhagem Celular Tumoral , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Mutação , Peptídeos/genética , Peptídeos/imunologia , Proteômica , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Int J Mol Sci ; 22(14)2021 Jul 07.
Artigo em Inglês | MEDLINE | ID: mdl-34298914

RESUMO

The loss of immune tolerance to fetal antigens may result in reproductive failure. The downregulated number and activity of T regulatory lymphocytes, which are critical for the establishment of immune tolerance to fetal antigens, during pregnancy may lead to miscarriage. The adoptive transfer of Tregs prevents fetal loss in abortion-prone mice. Recently, we demonstrated that the administration of tregitopes, which are short peptides found in human and mouse immunoglobulins (IgGs), decreased the incidence of abortions in female CBA/J mice mated with DBA/2J mice. Here, two non-IgG source peptides (SGS and LKD) that can potentially bind to the major histocompatibility complex II (MHC II) with high affinity and induce Treg expansion were designed in silico. The immune dysregulation-induced pregnancy failure mouse model was used to evaluate the effect of SGS and LKD on immune response and pregnancy outcome. The fetal death rate in the SGS-treated group was lower than that in the phosphate-buffered saline-treated group. SGS and LKD upregulated the splenic pool of Tregs and modulated the T-helper cell (Th1)/Th2-related cytokine response at the preimplantation stage. Additionally, SGS and LKD downregulated the expression of CD80 and MHC class II molecules in splenic CD11c+ antigen-presenting cells. Thus, SGS treatment can result in beneficial pregnancy outcomes. Additionally, SGS peptide-mediated immunomodulation can be a potential therapeutic strategy for immune dysregulation-induced pregnancy failure.


Assuntos
Aborto Espontâneo/imunologia , Células Apresentadoras de Antígenos/imunologia , Epitopos/imunologia , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/imunologia , Linfócitos T Reguladores/imunologia , Transferência Adotiva/métodos , Animais , Citocinas/imunologia , Modelos Animais de Doenças , Feminino , Antígenos de Histocompatibilidade Classe II/imunologia , Tolerância Imunológica/imunologia , Masculino , Camundongos , Camundongos Endogâmicos CBA , Gravidez , Resultado da Gravidez , Baço/imunologia , Células Th1/imunologia , Células Th2/imunologia
7.
Int J Mol Sci ; 22(12)2021 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-34207474

RESUMO

Chicken meat is often a major component of a modern diet. Allergy to chicken meat is relatively rare and occurs independently or in subjects allergic to ovalbumin (OVA). We examined the effect of adoptive transfer of OVA-CD4+ T cells on the immune response to OVA in mice fed chicken meat. Donor mice were injected intraperitoneally with 100 µg of OVA with Freund's adjuvant two times over a week, and CD4+ T cells were isolated from them and transferred to naïve mice (CD4+/OVA/ChM group), which were then provoked with OVA with FA and fed freeze-dried chicken meat for 14 days. The mice injected with OVA and fed chicken meat (OVA/ChM group), and sensitized (OVA group) and healthy (PBS group) mice served as controls. Humoral and cellular response to OVA was monitored over the study. The CD4+/OVA/ChM group had lowered levels of anti-OVA IgG and IgA, and total IgE. There were significant differences in CD4+, CD4+CD25+, and CD4+CD25+Foxp3+ T cells between groups. OVA stimulation decreased the splenocyte proliferation index and IFN-γ secretion in the CD4+/OVA/ChM group compared to the OVA group. IL-4 was increased in the OVA/ChM mice, which confirms allergenic potential of the egg-meat protein combination. Transfer of OVA-experienced CD4+ T cells ameliorated the negative immune response to OVA.


Assuntos
Transferência Adotiva , Alérgenos/imunologia , Linfócitos T CD4-Positivos/imunologia , Imunoglobulinas/imunologia , Ovalbumina/imunologia , Produtos Avícolas , Animais , Proteínas Aviárias/imunologia , Células Cultivadas , Galinhas , Feminino , Hipersensibilidade Alimentar/imunologia , Interferon gama/imunologia , Subpopulações de Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos BALB C
8.
Oncoimmunology ; 10(1): 1929726, 2021 05 25.
Artigo em Inglês | MEDLINE | ID: mdl-34104546

RESUMO

Adoptive cell therapy using TCR-engineered T cells (TCR-T cells) represents a promising strategy for treating relapsed and metastatic cancers. We previously established methods to identify neoantigen-specific TCRs based on patients' PBMCs. However, in clinical practice isolation of PBMCs from advanced-stage cancer patients proves to be difficult. In this study, we substituted blood-derived T cells for tumor-infiltrating lymphocytes (TILs) and used an HLA-matched cell line of antigen-presenting cells (APCs) to replace autologous dendritic cells. Somatic mutations were determined in head and neck squamous cell carcinoma resected from two patients. HLA-A*02:01-restricted neoantigen libraries were constructed and transferred into HLA-matched APCs for stimulation of patient TILs. TCRs were isolated from reactive TIL cultures and functionality was tested using TCR- T cells in vitro and in vivo. To exemplify the screening approach, we identified the targeted neoantigen leading to recognition of the minigene construct that stimulated the strongest TIL response. Neoantigen peptides were used to load MHC-tetramers for T cell isolation and a TCR was identified targeting the KIAA1429D1358E mutation. TCR-T cells were activated, exhibited cytotoxicity, and secreted cytokines in a dose-dependent manner, and only when stimulated with the mutant peptide. Furthermore, comparable to a neoantigen-specific TCR that was isolated from the patient's PBMCs, KIAA1429D1358E-specific TCR T cells destroyed human tumors in mice. The established protocol provides the required flexibility to methods striving to identify neoantigen-specific TCRs. By using an MHC-matched APC cell line and neoantigen-encoding minigene libraries, autologous TILs can be stimulated and screened when patient PBMCs and/or tumor material are not available anymore. Abbreviations: Head and neck squamous cell carcinoma (HNSCC); adoptive T cell therapy (ACT); T cell receptor (TCR); tumor-infiltrating lymphocytes (TIL); cytotoxic T lymphocyte (CTL); peripheral blood mononuclear cell (PBMC); dendritic cell (DC); antigen-presenting cells (APC).


Assuntos
Imunoterapia Adotiva , Receptores de Antígenos de Linfócitos T , Transferência Adotiva , Animais , Humanos , Linfócitos do Interstício Tumoral , Camundongos , Receptores de Antígenos de Linfócitos T/genética , Carcinoma de Células Escamosas de Cabeça e Pescoço/terapia
9.
Sci Immunol ; 6(60)2021 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-34145066

RESUMO

The nutrient-sensing mammalian target of rapamycin (mTOR) is integral to cell fate decisions after T cell activation. Sustained mTORC1 activity favors the generation of terminally differentiated effector T cells instead of follicular helper and memory T cells. This is particularly pertinent for T cell responses of older adults who have sustained mTORC1 activation despite dysfunctional lysosomes. Here, we show that lysosome-deficient T cells rely on late endosomes rather than lysosomes as an mTORC1 activation platform, where mTORC1 is activated by sensing cytosolic amino acids. T cells from older adults have an increased expression of the plasma membrane leucine transporter SLC7A5 to provide a cytosolic amino acid source. Hence, SLC7A5 and VPS39 deficiency (a member of the HOPS complex promoting early to late endosome conversion) substantially reduced mTORC1 activities in T cells from older but not young individuals. Late endosomal mTORC1 is independent of the negative-feedback loop involving mTORC1-induced inactivation of the transcription factor TFEB that controls expression of lysosomal genes. The resulting sustained mTORC1 activation impaired lysosome function and prevented lysosomal degradation of PD-1 in CD4+ T cells from older adults, thereby inhibiting their proliferative responses. VPS39 silencing of human T cells improved their expansion to pertussis and to SARS-CoV-2 peptides in vitro. Furthermore, adoptive transfer of CD4+ Vps39-deficient LCMV-specific SMARTA cells improved germinal center responses, CD8+ memory T cell generation, and recall responses to infection. Thus, curtailing late endosomal mTORC1 activity is a promising strategy to enhance T cell immunity.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , COVID-19/imunologia , Endossomos/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , SARS-CoV-2/metabolismo , Transdução de Sinais/genética , Transferência Adotiva/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Proteínas Relacionadas à Autofagia/deficiência , Proteínas Relacionadas à Autofagia/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , COVID-19/virologia , Células Cultivadas , Feminino , Proteína Forkhead Box O1/deficiência , Proteína Forkhead Box O1/genética , Voluntários Saudáveis , Humanos , Transportador 1 de Aminoácidos Neutros Grandes/metabolismo , Lisossomos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Transdução de Sinais/imunologia , Transfecção , Proteínas de Transporte Vesicular/deficiência , Proteínas de Transporte Vesicular/genética , Adulto Jovem
10.
Nat Immunol ; 22(7): 880-892, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34099917

RESUMO

Multidimensional single-cell analyses of T cells have fueled the debate about whether there is extensive plasticity or 'mixed' priming of helper T cell subsets in vivo. Here, we developed an experimental framework to probe the idea that the site of priming in the systemic immune compartment is a determinant of helper T cell-induced immunopathology in remote organs. By site-specific in vivo labeling of antigen-specific T cells in inguinal (i) or gut draining mesenteric (m) lymph nodes, we show that i-T cells and m-T cells isolated from the inflamed central nervous system (CNS) in a model of multiple sclerosis (MS) are distinct. i-T cells were Cxcr6+, and m-T cells expressed P2rx7. Notably, m-T cells infiltrated white matter, while i-T cells were also recruited to gray matter. Therefore, we propose that the definition of helper T cell subsets by their site of priming may guide an advanced understanding of helper T cell biology in health and disease.


Assuntos
Autoimunidade , Encéfalo/imunologia , Linhagem da Célula , Encefalomielite Autoimune Experimental/imunologia , Intestinos/imunologia , Pele/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Transferência Adotiva , Animais , Autoimunidade/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Sinalização do Cálcio , Líquido Cefalorraquidiano/imunologia , Líquido Cefalorraquidiano/metabolismo , Encefalomielite Autoimune Experimental/tratamento farmacológico , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/metabolismo , Cloridrato de Fingolimode/farmacologia , Perfilação da Expressão Gênica , Genes Codificadores dos Receptores de Linfócitos T , Células HEK293 , Humanos , Imunossupressores/farmacologia , Intestinos/efeitos dos fármacos , Microscopia Intravital , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia de Fluorescência , Esclerose Múltipla Recidivante-Remitente/genética , Esclerose Múltipla Recidivante-Remitente/imunologia , Esclerose Múltipla Recidivante-Remitente/metabolismo , Fenótipo , Estudos Prospectivos , RNA-Seq , Receptores CXCR6/genética , Receptores CXCR6/metabolismo , Receptores Purinérgicos P2X7/genética , Receptores Purinérgicos P2X7/metabolismo , Análise de Célula Única , Pele/efeitos dos fármacos , Pele/metabolismo , Linfócitos T Auxiliares-Indutores/efeitos dos fármacos , Linfócitos T Auxiliares-Indutores/metabolismo , Linfócitos T Auxiliares-Indutores/transplante , Transcriptoma
11.
Cancer Sci ; 112(9): 3437-3454, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34152672

RESUMO

Metastasis is the main cause of death in individuals with cancer. Immune checkpoint blockade (ICB) can potentially reverse CD8+ cytotoxic T lymphocytes (CTLs) dysfunction, leading to significant remission in multiple cancers. However, the mechanism underlying the development of CTL exhaustion during metastatic progression remains unclear. Here, we established an experimental pulmonary metastasis model with melanoma cells and discovered a critical role for melanoma-released exosomes in metastasis. Using genetic knockdown of nSMase2 and Rab27a, 2 key enzymes for exosome secretion, we showed that high levels of effector-like tumor-specific CD8+ T cells with transitory exhaustion, instead of terminal exhaustion, were observed in mice without exosomes; these cells showed limited inhibitory receptors and strong proliferation and cytotoxicity. Mechanistically, the immunosuppression of exosomes depends on exogenous PD-L1, which can be largely rescued by pretreatment with antibody blockade. Notably, we also found that exosomal PD-L1 acts as a promising predictive biomarker for ICB therapies during metastasis. Together, our findings suggest that exosomal PD-L1 may be a potential immunotherapy target, suggesting a new curative therapy for tumor metastasis.


Assuntos
Antígeno B7-H1/metabolismo , Exossomos/metabolismo , Tolerância Imunológica , Neoplasias Pulmonares/sangue , Neoplasias Pulmonares/secundário , Melanoma/metabolismo , Melanoma/patologia , Linfócitos T Citotóxicos/imunologia , Transferência Adotiva/métodos , Idoso , Animais , Antígeno B7-H1/antagonistas & inibidores , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Técnicas de Silenciamento de Genes , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Masculino , Melanoma/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Esfingomielina Fosfodiesterase/deficiência , Esfingomielina Fosfodiesterase/genética , Resultado do Tratamento , Proteínas rab27 de Ligação ao GTP/deficiência , Proteínas rab27 de Ligação ao GTP/genética
12.
Int J Mol Sci ; 22(10)2021 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-34065421

RESUMO

Dendritic cells (DCs) are unique immune cells that can link innate and adaptive immune responses and Immunometabolism greatly impacts their phenotype. Rapamycin is a macrolide compound that has immunosuppressant functions and is used to prevent graft loss in kidney transplantation. The current study evaluated the therapeutic potential of ex-vivo rapamycin treated DCs to protect kidneys in a mouse model of acute kidney injury (AKI). For the rapamycin single (S) treatment (Rapa-S-DC), Veh-DCs were treated with rapamycin (10 ng/mL) for 1 h before LPS. In contrast, rapamycin multiple (M) treatment (Rapa-M-DC) were exposed to 3 treatments over 7 days. Only multiple ex-vivo rapamycin treatments of DCs induced a persistent reprogramming of mitochondrial metabolism. These DCs had 18-fold more mitochondria, had almost 4-fold higher oxygen consumption rates, and produced more ATP compared to Veh-DCs (Veh treated control DCs). Pathway analysis showed IL10 signaling as a major contributing pathway to the altered immunophenotype after Rapamycin treatment compared to vehicle with significantly lower cytokines Tnfa, Il1b, and Il6, while regulators of mitochondrial content Pgc1a, Tfam, and Ho1 remained elevated. Critically, adoptive transfer of rapamycin-treated DCs to WT recipients 24 h before bilateral kidney ischemia significantly protected the kidneys from injury with a significant 3-fold improvement in kidney function. Last, the infusion of DCs containing higher mitochondria numbers (treated ex-vivo with healthy isolated mitochondria (10 µg/mL) one day before) also partially protected the kidneys from IRI. These studies demonstrate that pre-emptive infusion of ex-vivo reprogrammed DCs that have higher mitochondria content has therapeutic capacity to induce an anti-inflammatory regulatory phenotype to protect kidneys from injury.


Assuntos
Injúria Renal Aguda/tratamento farmacológico , Células Dendríticas/efeitos dos fármacos , Isquemia/tratamento farmacológico , Mitocôndrias/efeitos dos fármacos , Dinâmica Mitocondrial/efeitos dos fármacos , Traumatismo por Reperfusão/tratamento farmacológico , Sirolimo/farmacologia , Injúria Renal Aguda/metabolismo , Transferência Adotiva/métodos , Animais , Citocinas/metabolismo , Células Dendríticas/metabolismo , Células HEK293 , Humanos , Inflamação/metabolismo , Isquemia/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Traumatismo por Reperfusão/metabolismo , Transdução de Sinais/efeitos dos fármacos
13.
Nanotheranostics ; 5(4): 431-444, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33972919

RESUMO

Adoptive cellular therapy (ACT) is a potent strategy to boost the immune response against cancer. ACT is effective against blood cancers but faces challenges in treating solid tumors. A critical step for the success of ACT immunotherapy is to achieve efficient trafficking and persistence of T cells to solid tumors. Non-invasive tracking of the accumulation of adoptively transferred T cells to tumors would greatly accelerate development of more effective ACT strategies. We demonstrate the use of magnetic particle imaging (MPI) to non-invasively track ACT T cells in vivo in a mouse model of brain cancer. Magnetic labeling did not impair primary tumor-specific T cells in vitro, and MPI allowed the detection of labeled T cells in the brain after intravenous or intracerebroventricular administration. These results support the use of MPI to track adoptively transferred T cells and accelerate the development of ACT treatments for brain tumors and other cancers.


Assuntos
Transferência Adotiva , Neoplasias Encefálicas , Encéfalo , Rastreamento de Células , Nanopartículas de Magnetita/uso terapêutico , Linfócitos T , Animais , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/terapia , Camundongos , Camundongos Transgênicos , Linfócitos T/metabolismo , Linfócitos T/patologia , Linfócitos T/transplante
14.
Methods Mol Biol ; 2308: 139-150, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34057721

RESUMO

The bone marrow is the major hematopoietic organ, consisting of distinct microenvironmental niches for the production of hematopoietic cells. Advanced visualizing methods are required to define and better understand the interactions between stromal and hematopoietic cells. In this chapter, we describe an ex vivo whole-mount imaging technique of the bone marrow, which allows for a fast, high-quality, and three-dimensional visualization of different bone marrow components. We provide a guide for conducting adoptive transfer experiments of fluorescently labeled leukocytes and visualizing their location in the bone marrow with respect to the bone marrow vasculature. This method presents a quick, easy, and inexpensive approach to image the bone marrow in three dimensions.


Assuntos
Medula Óssea/irrigação sanguínea , Movimento Celular , Imageamento Tridimensional , Leucócitos/fisiologia , Microscopia Confocal , Microscopia de Fluorescência , Inclusão do Tecido , Transferência Adotiva , Animais , Microambiente Celular , Corantes Fluorescentes/metabolismo , Processamento de Imagem Assistida por Computador , Leucócitos/metabolismo , Camundongos , Microtomia
15.
Methods Mol Biol ; 2308: 163-176, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34057723

RESUMO

Decade-long survival of plasma cells in the bone marrow has long been a puzzling matter. To understand how plasma cells are maintained and supported by survival-niches to account for lifelong antibody production demands new intravital imaging techniques that are able to follow up a single cell and their interaction with other cell types in situ. We achieved to successfully establish longitudinal imaging of the bone marrow (LIMB) that is based on an implantable endoscopic device. In this chapter, basic approaches on how to investigate plasma cell-stroma interaction and surgical implantation procedures are introduced.


Assuntos
Células da Medula Óssea/fisiologia , Medula Óssea/fisiologia , Microambiente Celular , Processamento de Imagem Assistida por Computador , Microscopia Intravital , Microscopia de Fluorescência por Excitação Multifotônica , Plasmócitos/fisiologia , Transferência Adotiva , Animais , Medula Óssea/metabolismo , Células da Medula Óssea/metabolismo , Transplante de Medula Óssea , Separação Celular , Genes Reporter , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Camundongos Transgênicos , Plasmócitos/metabolismo , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo
16.
J Nutr Biochem ; 95: 108764, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-33964465

RESUMO

Obesity-associated inflammation in white adipose tissue (WAT) is a causal factor of systemic insulin resistance. To better understand how adipocytes regulate WAT inflammation, the present study generated chimeric mice in which inducible 6-phosphofructo-2-kinase was low, normal, or high in WAT while the expression of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (Pfkfb3) was normal in hematopoietic cells, and analyzed changes in high-fat diet (HFD)-induced WAT inflammation and systemic insulin resistance in the mice. Indicated by proinflammatory signaling and cytokine expression, the severity of HFD-induced WAT inflammation in WT â†’ Pfkfb3+/- mice, whose Pfkfb3 was disrupted in WAT adipocytes but not hematopoietic cells, was comparable with that in WT â†’ WT mice, whose Pfkfb3 was normal in all cells. In contrast, the severity of HFD-induced WAT inflammation in WT â†’ Adi-Tg mice, whose Pfkfb3 was over-expressed in WAT adipocytes but not hematopoietic cells, remained much lower than that in WT â†’ WT mice. Additionally, HFD-induced insulin resistance was correlated with the status of WAT inflammation and comparable between WT â†’ Pfkfb3+/- mice and WT â†’ WT mice, but was significantly lower in WT â†’ Adi-Tg mice than in WT â†’ WT mice. In vitro, palmitoleate decreased macrophage phosphorylation states of Jnk p46 and Nfkb p65 and potentiated the effect of interleukin 4 on suppressing macrophage proinflammatory activation. Taken together, these results suggest that the Pfkfb3 in adipocytes functions to suppress WAT inflammation. Moreover, the role played by adipocyte Pfkfb3 is attributable to, at least in part, palmitoleate promotion of macrophage anti-inflammatory activation.


Assuntos
Tecido Adiposo/metabolismo , Tecido Adiposo/patologia , Inflamação/metabolismo , Macrófagos/fisiologia , Fosfofrutoquinase-2/metabolismo , Adipócitos Brancos/metabolismo , Transferência Adotiva , Animais , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Dieta Hiperlipídica/efeitos adversos , Gorduras na Dieta/administração & dosagem , Gorduras na Dieta/farmacologia , Ácidos Graxos Monoinsaturados/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Intolerância à Glucose , Resistência à Insulina , Masculino , Camundongos , Fosfofrutoquinase-2/genética , Transdução de Sinais
17.
Nat Neurosci ; 24(6): 818-830, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33859437

RESUMO

Maternal immune activation (MIA) induced by lipopolysaccharides or polyinosinic:polycytidylic acid injections can induce behavioral abnormalities in adult mouse offspring. Here, we used the soluble tachyzoite antigen from Toxoplasma gondii, a parasite that infects approximately two billion people, to induce MIA in mice. The adult male offspring showed autism-relevant behaviors and abnormal brain microstructure, along with a pro-inflammatory T-cell immune profile in the periphery and upregulation of interleukin-6 in brain astrocytes. We show that adoptive transfer of regulatory T (Treg) cells largely reversed these MIA-induced phenotypes. Notably, pathogen-activated maternal Treg cells showed greater rescue efficacy than those from control donors. Single-cell RNA sequencing identified and characterized a unique group of pathogen-activated Treg cells that constitute 32.6% of the pathogen-activated maternal Treg population. Our study establishes a new preclinical parasite-mimicking MIA model and suggests therapeutic potential of adoptive Treg cell transfer in neuropsychiatric disorders associated with immune alterations.


Assuntos
Antígenos de Protozoários/toxicidade , Efeitos Tardios da Exposição Pré-Natal/imunologia , Efeitos Tardios da Exposição Pré-Natal/parasitologia , Interação Social , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/parasitologia , Transferência Adotiva/métodos , Animais , Antígenos de Protozoários/administração & dosagem , Feminino , Células HeLa , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Gravidez , Vocalização Animal/fisiologia
18.
J Virol ; 95(12)2021 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-33827945

RESUMO

Immune memory represents the most efficient defense against invasion and transmission of infectious pathogens. In contrast to memory T and B cells, the roles of innate immunity in recall responses remain inconclusive. In this study, we identified a novel mouse spleen NK cell subset expressing NKp46 and NKG2A induced by intranasal influenza virus infection. These memory NK cells specifically recognize N-linked glycosylation sites on influenza hemagglutinin (HA) protein. Different from memory-like NK cells reported previously, these NKp46+ NKG2A+ memory NK cells exhibited HA-specific silence of cytotoxicity but increase of gamma interferon (IFN-γ) response against influenza virus-infected cells, which could be reversed by pifithrin-µ, a p53-heat shock protein 70 (HSP70) signaling inhibitor. During recall responses, splenic NKp46+ NKG2A+ NK cells were recruited to infected lung and modulated viral clearance of virus and CD8+ T cell distribution, resulting in improved clinical outcomes. This long-lived NK memory bridges innate and adaptive immune memory response and promotes the homeostasis of local environment during recall response.IMPORTANCE In this study, we demonstrate a novel hemagglutinin (HA)-specific NKp46+ NKG2A+ NK cell subset induced by influenza A virus infection. These memory NK cells show virus-specific decreased cytotoxicity and increased gamma interferon (IFN-γ) on reencountering the same influenza virus antigen. In addition, they modulate host recall responses and CD8 T cell distribution, thus bridging the innate immune and adaptive immune responses during influenza virus infection.


Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Memória Imunológica , Vírus da Influenza A Subtipo H1N1/imunologia , Células Matadoras Naturais/imunologia , Infecções por Orthomyxoviridae/imunologia , Transferência Adotiva , Animais , Antígenos Ly/análise , Antígenos Ly/metabolismo , Benzotiazóis/farmacologia , Linfócitos T CD8-Positivos/imunologia , Técnicas de Cocultura , Citotoxicidade Imunológica , Células Dendríticas/imunologia , Vírus da Influenza A Subtipo H9N2/imunologia , Interferon gama/metabolismo , Células Matadoras Naturais/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Subfamília C de Receptores Semelhantes a Lectina de Células NK/análise , Receptor 1 Desencadeador da Citotoxicidade Natural/análise , Receptor 1 Desencadeador da Citotoxicidade Natural/metabolismo , Baço/citologia , Baço/imunologia , Tolueno/análogos & derivados , Tolueno/farmacologia
19.
Front Immunol ; 12: 626193, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33868244

RESUMO

Rheumatoid arthritis (RA) is a systemic and heterogeneous autoimmune disease with symmetrical polyarthritis as its critical clinical manifestation. The basic cause of autoimmune diseases is the loss of tolerance to self or harmless antigens. The loss or functional deficiency of key immune cells, regulatory T (Treg) cells, has been confirmed in human autoimmune diseases. The pathogenesis of RA is complex, and the dysfunction of Tregs is one of the proposed mechanisms underlying the breakdown of self-tolerance leading to the progression of RA. Treg cells are a vital component of peripheral immune tolerance, and the transcription factor Foxp3 plays a major immunosuppressive role. Clinical treatment for RA mainly utilizes drugs to alleviate the progression of disease and relieve disease activity, and the ideal treatment strategy should be to re-induce self-tolerance before obvious tissue injury. Treg cells are one of the ideal options. This review will introduce the classification, mechanism of action, and characteristics of Treg cells in RA, which provides insights into clinical RA treatment.


Assuntos
Artrite Reumatoide/imunologia , Autoimunidade , Tolerância a Antígenos Próprios , Linfócitos T Reguladores/imunologia , Transferência Adotiva , Animais , Antirreumáticos/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/metabolismo , Autoimunidade/efeitos dos fármacos , Citocinas/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Humanos , Mediadores da Inflamação/metabolismo , Fenótipo , Tolerância a Antígenos Próprios/efeitos dos fármacos , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/metabolismo , Linfócitos T Reguladores/transplante
20.
Front Immunol ; 12: 626172, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33912156

RESUMO

Regulatory T (Treg) cells are indispensable for immune homeostasis due to their roles in peripheral tolerance. As the master transcription factor of Treg cells, Forkhead box P3 (Foxp3) strongly regulates Treg function and plasticity. Because of this, considerable research efforts have been directed at elucidating the mechanisms controlling Foxp3 and its co-regulators. Such work is not only advancing our understanding on Treg cell biology, but also uncovering novel targets for clinical manipulation in autoimmune diseases, organ transplantation, and tumor therapies. Recently, many studies have explored the post-translational regulation of Foxp3, which have shown that acetylation, phosphorylation, glycosylation, methylation, and ubiquitination are important for determining Foxp3 function and plasticity. Additionally, some of these targets have been implicated to have great therapeutic values. In this review, we will discuss emerging evidence of post-translational regulations on Foxp3 in Treg cells and their exciting therapeutic applications.


Assuntos
Fatores de Transcrição Forkhead/metabolismo , Processamento de Proteína Pós-Traducional , Linfócitos T Reguladores/metabolismo , Transferência Adotiva , Animais , Humanos , Fatores Imunológicos/uso terapêutico , Fenótipo , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/transplante
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