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1.
PLoS Pathog ; 17(9): e1009818, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34529743

RESUMO

Cytotoxic lymphocytes produce granules armed with a set of 5 serine proteases (granzymes (Gzms)), which, together with the pore-forming protein (perforin), serve as a major defense against viral infections in humans. This granule-exocytosis pathway subsumes a well-established mechanism in which target cell death is induced upon perforin-mediated entry of Gzms and subsequent activation of various (apoptosis) pathways. In the past decade, however, a growing body of evidence demonstrated that Gzms also inhibit viral replication and potential reactivation in cell death-independent manners. For example, Gzms can induce proteolysis of viral or host cell proteins necessary for the viral entry, release, or intracellular trafficking, as well as augment pro-inflammatory antiviral cytokine response. In this review, we summarize current evidence for the noncytotoxic mechanisms and roles by which killer cells can use Gzms to combat viral infections, and we discuss the potential thereof for the development of novel therapies.


Assuntos
Granzimas/imunologia , Linfócitos T/imunologia , Viroses/imunologia , Humanos
2.
Front Immunol ; 12: 650788, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34220808

RESUMO

Numerous studies reported a small subpopulation of TCRαß+CD4-CD8- (double-negative) T cells that exert regulatory functions in the peripheral lymphocyte population. However, the origin of these double-negative T (DNT) cells is controversial. Some researchers reported that DNT cells originated from the thymus, and others argued that these cells are derived from peripheral immune induction. We report a possible mechanism for the induction of nonregulatory CD4+ T cells to become regulatory double-negative T (iDNT) cells in vitro. We found that immature bone marrow dendritic cells (CD86+MHC-II- DCs), rather than mature DCs (CD86+MHC-II+), induced high levels of iDNT cells. The addition of an anti-MHC-II antibody to the CD86+MHC-II+ DC group significantly increased induction. These iDNT cells promoted B cell apoptosis and inhibited B cell proliferation and plasma cell formation. A subgroup of iDNT cells expressed NKG2D. Compared to NKG2D- iDNT cells, NKG2D+ iDNT cells released more granzyme B to enhance B cell regulation. This enhancement may function via NKG2D ligands expressed on B cells following lipopolysaccharide stimulation. These results demonstrate that MHC-II impedes induction, and iDNT cells may be MHC independent. NKG2D expression on iDNT cells enhanced the regulatory function of these cells. Our findings elucidate one possible mechanism of the induction of peripheral immune tolerance and provide a potential treatment for chronic allograft rejection in the future.


Assuntos
Linfócitos B/imunologia , Antígenos CD4/imunologia , Antígenos CD8/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/imunologia , Linfócitos T/imunologia , Animais , Antígeno B7-2/genética , Antígeno B7-2/imunologia , Antígeno B7-2/metabolismo , Antígenos CD4/genética , Antígenos CD4/metabolismo , Antígenos CD8/genética , Antígenos CD8/metabolismo , Células Cultivadas , Citocinas/genética , Citocinas/imunologia , Citocinas/metabolismo , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Expressão Gênica/imunologia , Granzimas/genética , Granzimas/imunologia , Granzimas/metabolismo , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe II/imunologia , Antígenos de Histocompatibilidade Classe II/metabolismo , Masculino , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Imunológicos , Subfamília K de Receptores Semelhantes a Lectina de Células NK/genética , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Linfócitos T/metabolismo
3.
Front Immunol ; 12: 689302, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34177946

RESUMO

Salmon Gill Poxvirus Disease (SGPVD) has emerged as a cause of acute mortality in Atlantic salmon (Salmo salar L.) presmolts in Norwegian aquaculture. The clinical phase of the disease is associated with apoptotic cell death in the gill epithelium causing acute respiratory distress, followed by proliferative changes in the regenerating gill in the period after the disease outbreak. In an experimental SGPV challenge trial published in 2020, acute disease was only seen in fish injected with hydrocortisone 24 h prior to infection. SGPV-mediated mortality in the hydrocortisone-injected group was associated with more extensive gill pathology and higher SGPV levels compared to the group infected with SGPV only. In this study based on the same trial, SGPV gene expression and the innate and adaptive antiviral immune response was monitored in gills and spleen in the presence and absence of hydrocortisone. Whereas most SGPV genes were induced from day 3 along with the interferon-regulated innate immune response in gills, the putative SGPV virulence genes of the B22R family were expressed already one day after SGPV exposure, indicating a potential role as early markers of SGPV infection. In gills of the hydrocortisone-injected fish infected with SGPV, MX expression was delayed until day 10, and then expression skyrocketed along with the viral peak, gill pathology and mortality occurring from day 14. A similar expression pattern was observed for Interferon gamma (IFNγ) and granzyme A (GzmA) in the gills, indicating a role of acute cytotoxic cell activity in SGPVD. Duplex in situ hybridization demonstrated effects of hydrocortisone on the number and localization of GzmA-containing cells, and colocalization with SGPV infected cells in the gill. SGPV was generally not detected in spleen, and gill infection did not induce any corresponding systemic immune activity in the absence of stress hormone injection. However, in fish injected with hydrocortisone, IFNγ and GzmA gene expression was induced in spleen in the days prior to acute mortality. These data indicate that suppressed mucosal immune response in the gills and the late triggered systemic immune response in the spleen following hormonal stress induction may be the key to the onset of clinical SGPVD.


Assuntos
Anti-Inflamatórios/farmacologia , Doenças dos Peixes/imunologia , Hidrocortisona/farmacologia , Imunidade nas Mucosas/efeitos dos fármacos , Infecções por Poxviridae/imunologia , Salmo salar/imunologia , Animais , Antígenos CD4/genética , Antígenos CD4/imunologia , Antígenos CD8/genética , Antígenos CD8/imunologia , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Brânquias/imunologia , Brânquias/virologia , Granzimas/genética , Granzimas/imunologia , Interações Hospedeiro-Patógeno , Interferon gama/genética , Interferon gama/imunologia , Membrana Mucosa/imunologia , Poxviridae/genética , Salmo salar/genética , Salmo salar/virologia
4.
Int J Mol Sci ; 22(11)2021 May 26.
Artigo em Inglês | MEDLINE | ID: mdl-34073458

RESUMO

Cytotoxic CD4+ T cells (CD4 CTL) are terminally differentiated T helper cells that contribute to autoimmune diseases, such as multiple sclerosis. We developed a novel triple co-culture transwell assay to study mutual interactions between CD4 CTL, conventional TH cells, and regulatory T cells (Tregs) simultaneously. We show that, while CD4 CTL are resistant to suppression by Tregs in vitro, the conditioned medium of CD4 CTL accentuates the suppressive phenotype of Tregs by upregulating IL-10, Granzyme B, CTLA-4, and PD-1. We demonstrate that CD4 CTL conditioned medium skews memory TH cells to a TH17 phenotype, suggesting that the CD4 CTL induce bystander polarization. In our triple co-culture assay, the CD4 CTL secretome promotes the proliferation of TH cells, even in the presence of Tregs. However, when cell-cell contact is established between CD4 CTL and TH cells, the proliferation of TH cells is no longer increased and Treg-mediated suppression is restored. Taken together, our results suggest that when TH cells acquire cytotoxic properties, these Treg-resistant CD4 CTL affect the proliferation and phenotype of conventional TH cells in their vicinity. By creating such a pro-inflammatory microenvironment, CD4 CTL may favor their own persistence and expansion, and that of other potentially pathogenic TH cells, thereby contributing to pathogenic responses in autoimmune disorders.


Assuntos
Doenças Autoimunes/imunologia , Proliferação de Células , Linfócitos T Reguladores/imunologia , Células Th17/imunologia , Adulto , Antígeno CTLA-4/imunologia , Feminino , Granzimas/imunologia , Humanos , Interleucina-10/imunologia , Masculino , Pessoa de Meia-Idade , Receptor de Morte Celular Programada 1/imunologia , Linfócitos T Reguladores/citologia , Células Th17/citologia
5.
Front Immunol ; 12: 677707, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34017346

RESUMO

Granzymes are a family of serine proteases stored in granules inside cytotoxic cells of the immune system. Granzyme K (GrK) has been only limitedly characterized and knowledge on its molecular functions is emerging. Traditionally GrK is described as a granule-secreted, pro-apoptotic serine protease. However, accumulating evidence is redefining the functions of GrK by the discovery of novel intracellular (e.g. cytotoxicity, inhibition of viral replication) and extracellular roles (e.g. endothelial activation and modulation of a pro-inflammatory immune cytokine response). Moreover, elevated GrK levels are associated with disease, including viral and bacterial infections, airway inflammation and thermal injury. This review aims to summarize and discuss the current knowledge of i) intracellular and extracellular GrK activity, ii) cytotoxic and non-cytotoxic GrK functioning, iii) the role of GrK in disease, and iv) GrK as a potential therapeutic target.


Assuntos
Espaço Extracelular/imunologia , Granzimas/imunologia , Granzimas/metabolismo , Espaço Intracelular/imunologia , Animais , Infecções Bacterianas/tratamento farmacológico , Infecções Bacterianas/imunologia , Infecções Bacterianas/metabolismo , Citocinas/metabolismo , Citotoxicidade Imunológica , Células Endoteliais/imunologia , Células Endoteliais/metabolismo , Espaço Extracelular/metabolismo , Granzimas/antagonistas & inibidores , Humanos , Espaço Intracelular/metabolismo , Pneumopatias/tratamento farmacológico , Pneumopatias/imunologia , Pneumopatias/metabolismo , Terapia de Alvo Molecular/métodos , Inibidores de Serino Proteinase/uso terapêutico , Resultado do Tratamento , Viroses/tratamento farmacológico , Viroses/imunologia , Viroses/metabolismo
6.
Front Immunol ; 12: 578548, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33815355

RESUMO

Objectives: To explore the potential role of CD3+CD8+CD161high TCRVα7.2+ mucosal-associated invariant T (MAIT) cells in the pathogenesis of primary biliary cholangitis (PBC). Methods: We enrolled 55 patients with PBC, 69 healthy controls (HCs), and 8 patients with hepatic hemangioma. Circulating MAIT cells and their chemokine receptor profiles and cytokine production were quantified using flow cytometry. Liver-resident MAIT cells were examined by immunofluorescence staining. CXCL12-mediated chemotaxis of MAIT cells was measured using a transwell migration assay. Plasma interleukin (IL)-18 was measured using ELISA, and cytokine production in IL-18-stimulated MAIT cells was detected using flow cytometry. Result: Peripheral MAIT cells were found to be significantly lower in patients with PBC (3.0 ± 3.2% vs. 9.4 ± 8.0%, p < 0.01) and negatively correlated with alkaline phosphatase (ALP) levels (r = -0.3209, p < 0.05). Liver immunofluorescence staining suggested that MAIT cells might accumulate in PBC liver. MAIT cells from patients with PBC expressed higher levels of CXCR4 (84.8 ± 18.0% vs. 58.7 ± 11.4%, p < 0.01), and the expression of CXCL12 was higher in PBC liver. CXCL12 promoted MAIT cell chemotaxis (70.4 ± 6.8% vs. 52.2 ± 3.5%, p < 0.01), which was attenuated by CXCR4 antagonist. MAIT cells from PBC produced significantly more interferon-γ (IFN-γ) (88.3 ± 4.2% vs. 64.2 ± 10.1%, p < 0.01), tumor necrosis factor-α (TNF-α) (93.0 ± 1.1% vs. 80.1 ± 5.3%, p < 0.01), Granzyme B (89.3 ± 3.3% vs. 72.1 ± 7.0%, p < 0.01), and perforin (46.8 ± 6.6% vs. 34.8 ± 7.7%, p < 0.05). MAIT cells from PBC expressed higher levels of IL18-Rα (83.8 ± 10.2% vs. 58.3 ± 8.7%, p < 0.01). Plasma IL-18 was more abundant in patients with PBC (286.8 ± 75.7 pg/ml vs. 132.9 ± 78.1 pg/ml, p < 0.01). IL-18 promoted IFN-γ production in MAIT cells (74.9 ± 6.6% vs. 54.7 ± 6.7%, p < 0.01), which was partially attenuated by blocking IL-18R (68.6 ± 8.3% vs. 43.5 ± 4.2%, p < 0.01). Conclusion: Mucosal-associated invariant T cells from patients with PBC accumulated in the liver via CXCL12-CXCR4-mediated chemotaxis, produced pro-inflammatory cytokines, and contributed to portal inflammation, which was potentially mediated by elevated IL-18. Targeting MAIT cells might be a therapeutic approach for PBC.


Assuntos
Quimiocina CXCL12/imunologia , Cirrose Hepática Biliar/imunologia , Fígado/imunologia , Células T Invariantes Associadas à Mucosa/imunologia , Receptores CXCR4/imunologia , Adulto , Fosfatase Alcalina/imunologia , Fosfatase Alcalina/metabolismo , Quimiocina CXCL12/metabolismo , Quimiotaxia/imunologia , Feminino , Granzimas/imunologia , Granzimas/metabolismo , Humanos , Interferon gama/imunologia , Interferon gama/metabolismo , Interleucina-18/imunologia , Interleucina-18/metabolismo , Fígado/metabolismo , Cirrose Hepática Biliar/sangue , Cirrose Hepática Biliar/metabolismo , Contagem de Linfócitos , Masculino , Pessoa de Meia-Idade , Células T Invariantes Associadas à Mucosa/metabolismo , Perforina/imunologia , Perforina/metabolismo , Receptores CXCR4/metabolismo , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismo
7.
Front Immunol ; 12: 670324, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33868318

RESUMO

Granzyme B is a renowned effector molecule primarily utilized by CTLs and NK cells against ill-defined and/or transformed cells during immunosurveillance. The overall expression of granzyme B within tumor microenvironment has been well-established as a prognostic marker indicative of priming immunity for a long time. Until recent years, increasing immunosuppressive effects of granzyme B are unveiled in the setting of different immunological context. The accumulative evidence confounded the roles of granzyme B in immune responses, thereby arousing great interests in characterizing detailed feature of granzyme B-positive niche. In this paper, the granzyme B-related regulatory effects of major suppressor cells as well as the tumor microenvironment that defines such functionalities were longitudinally summarized and discussed. Multiplex networks were built upon the interactions among different transcriptional factors, cytokines, and chemokines that regarded to the initiation and regulation of granzyme B-mediated immunosuppression. The conclusions and prospect may facilitate better interpretations of the clinical significance of granzyme B, guiding the rational development of therapeutic regimen and diagnostic probes for anti-tumor purposes.


Assuntos
Granzimas/imunologia , Transdução de Sinais/imunologia , Evasão Tumoral/imunologia , Microambiente Tumoral/imunologia , Animais , Humanos
8.
Clin Exp Immunol ; 205(1): 53-62, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33735518

RESUMO

High expression of the inhibitory receptor programmed cell death ligand 1 (PD-L1) on tumor cells and tumor stromal cells have been found to play a key role in tumor immune evasion in several human malignancies. However, the expression of PD-L1 on bone marrow mesenchymal stem cells (BMSCs) and whether the programmed cell death 1 (PD-1)/PD-L1 signal pathway is involved in the BMSCs versus T cell immune response in multiple myeloma (MM) remains poorly defined. In this study, we explored the expression of PD-L1 on BMSCs from newly diagnosed MM (NDMM) patients and the role of PD-1/PD-L1 pathway in BMSC-mediated regulation of CD8+ T cells. The data showed that the expression of PD-L1 on BMSCs in NDMM patients was significantly increased compared to that in normal controls (NC) (18·81 ± 1·61 versus 2·78± 0·70%; P < 0·001). Furthermore, the PD-1 expression on CD8+ T cells with NDMM patients was significantly higher than that in normal controls (43·22 ± 2·98 versus 20·71 ± 1·08%; P < 0·001). However, there was no significant difference in PD-1 expression of CD4+ T cells and natural killer (NK) cells between the NDMM and NC groups. Additionally, the co-culture assays revealed that BMSCs significantly suppressed CD8+ T cell function. However, the PD-L1 inhibitor effectively reversed BMSC-mediated suppression in CD8+ T cells. We also found that the combination of PD-L1 inhibitor and pomalidomide can further enhance the killing effect of CD8+ T cells on MM cells. In summary, our findings demonstrated that BMSCs in patients with MM may induce apoptosis of CD8+ T cells through the PD-1/PD-L1 axis and inhibit the release of perforin and granzyme B from CD8+ T cells to promote the immune escape of MM.


Assuntos
Antígeno B7-H1/imunologia , Medula Óssea/imunologia , Linfócitos T CD8-Positivos/imunologia , Imunidade/imunologia , Células-Tronco Mesenquimais/imunologia , Mieloma Múltiplo/imunologia , Receptor de Morte Celular Programada 1/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos T CD4-Positivos/imunologia , Feminino , Granzimas/imunologia , Humanos , Células Matadoras Naturais/imunologia , Masculino , Pessoa de Meia-Idade , Perforina/imunologia , Transdução de Sinais/imunologia , Talidomida/análogos & derivados , Talidomida/imunologia , Evasão Tumoral/imunologia
9.
Cancer Immunol Immunother ; 70(11): 3137-3154, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33779796

RESUMO

BACKGROUND: Skin melanoma is a highly immunogenic cancer. The intratumoral immune cytolytic activity (CYT) reflects the ability of cytotoxic T and NK cells to eliminate cancer cells, and is associated with improved patient survival. Despite the enthusiastic clinical results seen in advanced-stage metastatic melanoma patients treated with immune checkpoint inhibitors, a subgroup of them will later relapse and develop acquired resistance. We questioned whether CYT associates with different genomic profiles and thus, patient outcome, in skin melanoma. METHODS: We explored the TCGA-SKCM dataset and stratified patients to distinct subgroups of cytolytic activity. The tumor immune contexture, somatic mutations and recurrent copy number aberrations were calculated using quanTIseq, MutSigCV and GISTIC2. Chromothriptic events were explored using CTLPScanner and cancer neoepitopes were predicted with antigen garnish. Each tumor's immunophenoscore was calculated using Immunophenogram. Mutational signatures and kataegis were explored using SigProfiler and compared to the known single or doublet base substitution signatures from COSMIC. RESULTS: Metastatic skin melanomas had significantly higher CYT levels compared to primary tumors. We assessed enrichment for immune-related gene sets within CYT-high tumors, whereas, CYT-low tumors were enriched for non-immune related gene sets. In addition, distinct mutational and neoantigen loads, primarily composed of C > T transitions, along with specific types of copy number aberrations, characterized each cytolytic subgroup. We found a broader pattern of chromothripsis across CYT-low tumors, where chromosomal regions harboring chromothriptic events, contained a higher number of cancer genes. SBS7a/b, SBS5 and SBS1 were the most prevalent mutational signatures across both cytolytic subgroups, but SBS1 differed significantly between them. SBS7a/b was mutually exclusive with SBS5 and SBS1 in both CYT subgroups. CYT-high patients had markedly higher immunophenoscore, suggesting that they should display a clinical benefit upon treatment with immune checkpoint inhibition therapy, compared to CYT-low patients. CONCLUSIONS: Overall, our data highlight the existence of distinct genomic features across cytolytic subgroups in skin melanoma, which might affect the patients' relapse rate or their acquisition of resistance to immune checkpoint inhibition therapies.


Assuntos
Citotoxicidade Imunológica/imunologia , Melanoma/genética , Melanoma/imunologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/imunologia , Granzimas/imunologia , Humanos , Células Matadoras Naturais/imunologia , Mutação , Perforina/imunologia , Linfócitos T Citotóxicos/imunologia
10.
J Immunother Cancer ; 9(3)2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33707314

RESUMO

While vaccines directed against the SARS-CoV-2 spike protein will have varying degrees of effectiveness in preventing SARS-CoV-2 infections, the severity of infection will be determined by multiple host factors including the ability of immune cells to lyse virus-infected cells. This review will discuss the complexity of both adaptive and innate immunomes and how a flow-based assay can detect up to 158 distinct cell subsets in the periphery. This assay has been employed to show the effect of age on differences in specific immune cell subsets, and the differences in the immunome between healthy donors and age-matched cancer patients. Also reviewed are the numerous soluble factors, in addition to cytokines, that may vary in the pathogenesis of SARS-CoV-2 infections and may also be employed to help define the effectiveness of a given vaccine or other antiviral agents. Various steroids have been employed in the management of autoimmune adverse events in cancer patients receiving immunotherapeutics and may be employed in the management of SARS-CoV-2 infections. The influence of steroids on multiple immune cells subsets will also be discussed.


Assuntos
Imunidade Adaptativa/imunologia , Linfócitos B/imunologia , COVID-19/imunologia , Células Dendríticas/imunologia , Imunidade Inata/imunologia , Células Matadoras Naturais/imunologia , Neoplasias/imunologia , Linfócitos T/imunologia , Fatores Etários , Antígeno B7-H1/imunologia , Ligante de CD40/imunologia , COVID-19/prevenção & controle , Vacinas contra COVID-19/uso terapêutico , Citocinas/imunologia , Suscetibilidade a Doenças , Glucocorticoides/uso terapêutico , Granzimas/imunologia , Humanos , Inibidores de Checkpoint Imunológico/uso terapêutico , Imunossenescência/imunologia , Células Supressoras Mieloides/imunologia , Neoplasias/tratamento farmacológico , Receptor de Morte Celular Programada 1/imunologia , Proteoma , SARS-CoV-2 , Índice de Gravidade de Doença , Subpopulações de Linfócitos T/imunologia , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/imunologia
11.
Mol Cell ; 81(7): 1469-1483.e8, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33609448

RESUMO

We demonstrate that DNA hypomethylating agent (HMA) treatment can directly modulate the anti-tumor response and effector function of CD8+ T cells. In vivo HMA treatment promotes CD8+ T cell tumor infiltration and suppresses tumor growth via CD8+ T cell-dependent activity. Ex vivo, HMAs enhance primary human CD8+ T cell activation markers, effector cytokine production, and anti-tumor cytolytic activity. Epigenomic and transcriptomic profiling shows that HMAs vastly regulate T cell activation-related transcriptional networks, culminating with over-activation of NFATc1 short isoforms. Mechanistically, demethylation of an intragenic CpG island immediately downstream to the 3' UTR of the short isoform was associated with antisense transcription and alternative polyadenylation of NFATc1 short isoforms. High-dimensional single-cell mass cytometry analyses reveal a selective effect of HMAs on a subset of human CD8+ T cell subpopulations, increasing both the number and abundance of a granzyme Bhigh, perforinhigh effector subpopulation. Overall, our findings support the use of HMAs as a therapeutic strategy to boost anti-tumor immune response.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Ilhas de CpG/imunologia , Metilação de DNA/efeitos dos fármacos , Decitabina/farmacologia , Granzimas/imunologia , Ativação Linfocitária/efeitos dos fármacos , Metilação de DNA/imunologia , Humanos , Fatores de Transcrição NFATC/imunologia , Perforina/imunologia
12.
PLoS Negl Trop Dis ; 15(2): e0009059, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33539379

RESUMO

BACKGROUND: Signs of senescence and the late stages of differentiation associated with the more severe forms of Chagas disease have been described in the Trypanosoma cruzi antigen-specific CD4+ T-cell population. However, the mechanisms involved in these functions are not fully known. To date, little is known about the possible impact of benznidazole treatment on the T. cruzi-specific functional response of CD4+ T cells. METHODOLOGY/PRINCIPAL FINDINGS: The functional capacity of CD4+ T cells was analyzed by cytometric assays in chronic Chagas disease patients, with indeterminate form (IND) and cardiac alterations (CCC) (25 and 15, respectively) before and after benznidazole treatment. An increase in the multifunctional capacity (expression of IFN-γ, IL-2, TNF-α, perforin and/or granzyme B) of the antigen-specific CD4+ T cells was observed in indeterminate versus cardiac patients, which was associated with the reduced coexpression of inhibitory receptors (2B4, CD160, CTLA-4, PD-1 and/or TIM-3). The functional profile of these cells shows statistically significant differences between IND and CCC (p<0.001), with a higher proportion of CD4+ T cells coexpressing 2 and 3 molecules in IND (54.4% versus 23.1% and 4.1% versus 2.4%, respectively). A significant decrease in the frequencies of CD4+ T cells that coexpress 2, 3 and 4 inhibitory receptors was observed in IND after 24-48 months of treatment (p<0.05, p<0.01 and p<0.05, respectively), which was associated with an increase in antigen-specific multifunctional activity. The IND group showed, at 9-12 months after treatment, an increase in the CD4+ T cell subset coproducing three molecules, which were mainly granzyme B+, perforin+ and IFN-γ+ (1.4% versus 4.5%). CONCLUSIONS/SIGNIFICANCE: A CD4+ T cell dysfunctional process was detected in chronic Chagas disease patients, being more exacerbated in those patients with cardiac symptoms. After short-term benznidazole treatment (9-12 months), indeterminate patients showed a significant increase in the frequency of multifunctional antigen-specific CD4+ T cells.


Assuntos
Antiprotozoários/administração & dosagem , Linfócitos T CD4-Positivos/imunologia , Doença de Chagas/tratamento farmacológico , Nitroimidazóis/administração & dosagem , Trypanosoma cruzi/efeitos dos fármacos , Adulto , Anticorpos Antiprotozoários/imunologia , Doença de Chagas/imunologia , Doença de Chagas/parasitologia , Feminino , Granzimas/imunologia , Humanos , Interferon gama/imunologia , Masculino , Pessoa de Meia-Idade , Perforina/imunologia , Espanha , Trypanosoma cruzi/genética , Trypanosoma cruzi/imunologia , Adulto Jovem
13.
Methods Mol Biol ; 2270: 203-216, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33479900

RESUMO

Granzyme B (GZMB)-expressing B cells inhibit CD4+ T-lymphocyte proliferation in a contact- and GZMB-dependent manner, through degradation of TCR zeta or induction of T-cell apoptosis. This regulatory B-cell population is present in human healthy individuals and represents about 1% of circulating B cells. Their small proportion requires the development of expansion methods to enable their study and envision clinical applications. We describe here how to expand GZMB-expressing B cells to obtain more than 90% of highly purified GZMB+ B cells, and the protocol of B/T cells coculture for the evaluation of the suppressive function of the GZMB+ B-cell population.


Assuntos
Linfócitos B Reguladores/imunologia , Técnicas de Cultura de Células/métodos , Granzimas/isolamento & purificação , Apoptose/imunologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Linfócitos B Reguladores/metabolismo , Proliferação de Células , Granzimas/imunologia , Granzimas/metabolismo , Humanos , Ativação Linfocitária , Linfócitos T Citotóxicos/imunologia
14.
Eur J Immunol ; 51(2): 483-486, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-32949467

RESUMO

Brain CD8+ CD69+ tissue-resident memory T (TRM ) cells comprise a CD20dim subset, which is proportionally larger in CD103-negative TRM cells. In multiple sclerosis (MS) lesions, CD20dim TRM -cell proportions are increased. CD20-expression is associated with higher levels of CXCR6, Ki-67, and granzyme B, supporting CD20dim TRM cells as a relevant subset in MS.


Assuntos
Antígenos CD20/imunologia , Linfócitos T CD8-Positivos/imunologia , Memória Imunológica/imunologia , Esclerose Múltipla/imunologia , Esclerose Múltipla/patologia , Substância Branca/imunologia , Substância Branca/patologia , Granzimas/imunologia , Humanos , Antígeno Ki-67/imunologia , Receptores CXCR6/imunologia
15.
J Obstet Gynaecol ; 41(1): 94-99, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32608278

RESUMO

The chronic course of endometriosis suggests that the immune system may play a role in its aetiology. There may be resistance to cell lysis, as well as an immune defect underlying endometriosis. Granzyme B is a serine protease that is secreted by Natural Killer (NK) cells and cytotoxic T lymphocytes during a cellular immune response and can induce apoptosis. The aim of this study was to evaluate the relationship between both Granzyme B levels and Granzyme B gene polymorphisms in endometriosis patients. Women between the ages of 20 - 45 were included in the study. The patients were divided into two groups: those diagnosed with endometriosis and those who had not been diagnosed with endometriosis. In the blood samples, Granzyme B gene polymorphisms and serum levels of Granzyme B were studied. There was no difference between the groups in terms of median Granzyme B levels and the presence of AA, AG, and GG genotypes. There was a difference in median granzyme levels for the control group; the GG genotype was found at a lower frequency. The immune defect within endometriosis-related immune cells may not be exclusively due to Granzyme B. Other mediators that are secreted from immune cells may have additive effects.IMPACT STATEMENTWhat is already known on this subject? NK cells are cytotoxic and inhibit the implantation of autologous endometrial cells that are spilled into the peritoneum by retrograde menstruation. Thus, a reduction in NK cell activity may facilitate the progression of endometriosis. The literature review reveals that there are studies suggesting that NK cell activity may be insufficient in endometriosis. Granzyme B is a serine protease that is secreted by NK cells and cytotoxic T lymphocytes during a cellular immune response.What do the results of this study add? Granzyme B is one of the cytotoxic granules in NK and cytotoxic T lymphocyte cells and its genetic polymorphisms were tested in endometriosis. We found that median Granzyme B levels were significantly different in patients with the GG genotype in the control group, compared to those with the AA and AG genotype. However, this difference was not detected between the control and endometriosis groups.What are the implications of these findings for clinical practice and/or further research? Our results contribute to uncovering the pathogenesis of endometriosis since there are no previous studies in the literature regarding this topic. Although we did not find a difference, our results will inform further studies made on this topic. Studies with different molecules and an increased number of patients are needed. The immune defect of endometriosis may not be due exclusively to Granzyme B. Other mediators that are secreted from immune cells may have mutual effects and interactions.


Assuntos
Endometriose/genética , Endometriose/imunologia , Granzimas/sangue , Imunidade Celular/genética , Polimorfismo Genético/imunologia , Adulto , Endometriose/sangue , Endométrio/enzimologia , Endométrio/imunologia , Feminino , Genótipo , Granzimas/imunologia , Humanos , Células Matadoras Naturais/enzimologia , Pessoa de Meia-Idade , Adulto Jovem
16.
Anticancer Agents Med Chem ; 21(6): 738-746, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32723258

RESUMO

BACKGROUND: The antigen HCA587 (also known as MAGE-C2), which is considered a cancer-testis antigen, exhibits upregulated expression in a wide range of malignant tumors with unique immunological properties, and may thus serve as a promising target for tumor immunotherapy. OBJECTIVE: The study aimed to explore the antitumor effect of the HCA587 protein vaccine and the response of humoral and cell-mediated immunity. METHODS: The HCA587 protein vaccine was formulated with adjuvants CpG and ISCOM. B16 melanoma cells were subcutaneously inoculated to C57BL/6 mice, followed by treatment with HCA587 protein vaccine subcutaneously. Mouse survival was monitored daily, and tumor volume was measured every 2 to 3 days. The tumor sizes, survival time and immune cells in tumor tissues were detected. And the vital immune cell subset and effector molecules were explored. RESULTS: After treatment with HCA587 protein vaccine, the vaccination elicited significant immune responses, which delayed tumor growth and improved animal survival. The vaccination increased the proportion of CD4+ T cells expressing IFN-γ and granzyme B in tumor tissues. The depletion of CD4+T cells resulted in an almost complete abrogation of the antitumor effect of the vaccination, suggesting that the antitumor efficacy was mediated by CD4+ T cells. In addition, knockout of IFN-γ resulted in a decrease in granzyme B levels, which were secreted by CD4+ T cells, and the antitumor effect was also significantly attenuated. CONCLUSION: The HCA587 protein vaccine may increase the levels of granzyme B expressed by CD4+ T cells, and this increase is dependent on IFN-γ, and the vaccine resulted in a specific tumor immune response and subsequent eradication of the tumor.


Assuntos
Antígenos de Neoplasias/imunologia , Linfócitos T CD4-Positivos/imunologia , Vacinas Anticâncer/imunologia , Granzimas/imunologia , Melanoma Experimental/prevenção & controle , Proteínas de Neoplasias/imunologia , Adjuvantes Imunológicos , Animais , Composição de Medicamentos , Regulação da Expressão Gênica/imunologia , Granzimas/genética , Humanos , Imunidade , Imunoterapia , Interferon gama/genética , Interferon gama/imunologia , Masculino , Melanoma Experimental/genética , Melanoma Experimental/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Experimentais
17.
Front Immunol ; 11: 584423, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33262762

RESUMO

Immune checkpoint therapy (ICT) results in durable responses in individuals with some cancers, but not all patients respond to treatment. ICT improves CD8+ cytotoxic T lymphocyte (CTL) function, but changes in tumor antigen-specific CTLs post-ICT that correlate with successful responses have not been well characterized. Here, we studied murine tumor models with dichotomous responses to ICT. We tracked tumor antigen-specific CTL frequencies and phenotype before and after ICT in responding and non-responding animals. Tumor antigen-specific CTLs increased within tumor and draining lymph nodes after ICT, and exhibited an effector memory-like phenotype, expressing IL-7R (CD127), KLRG1, T-bet, and granzyme B. Responding tumors exhibited higher infiltration of effector memory tumor antigen-specific CTLs, but lower frequencies of regulatory T cells compared to non-responders. Tumor antigen-specific CTLs persisted in responding animals and formed memory responses against tumor antigens. Our results suggest that increased effector memory tumor antigen-specific CTLs, in the presence of reduced immunosuppression within tumors is part of a successful ICT response. Temporal and nuanced analysis of T cell subsets provides a potential new source of immune based biomarkers for response to ICT.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Inibidores de Checkpoint Imunológico/imunologia , Memória Imunológica/imunologia , Animais , Antígenos de Neoplasias/imunologia , Biomarcadores/metabolismo , Linhagem Celular Tumoral , Granzimas/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Linfócitos T Citotóxicos/imunologia , Linfócitos T Reguladores/imunologia
18.
Front Immunol ; 11: 587581, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33262766

RESUMO

Inflammation is strictly interconnected to anti-inflammatory mechanisms to maintain tissue homeostasis. The disruption of immune homeostasis can lead to acute and chronic inflammatory diseases, as cardiovascular, pulmonary, metabolic diseases and cancer. The knowledge of the mechanisms involved in the development and progression of these pathological conditions is important to find effective therapies. Granzyme B (GrB) is a serine protease produced by a variety of immune, non-immune and tumor cells. Apoptotic intracellular and multiple extracellular functions of GrB have been recently identified. Its capability of cleaving extracellular matrix (ECM) components, cytokines, cell receptors and clotting proteins, revealed GrB as a potential multifunctional pro-inflammatory molecule with the capability of contributing to the pathogenesis of different inflammatory conditions, including inflammaging, acute and chronic inflammatory diseases and cancer. Here we give an overview of recent data concerning GrB activity on multiple targets, potentially allowing this enzyme to regulate a wide range of crucial biological processes that play a role in the development, progression and/or severity of inflammatory diseases. We focus our attention on the promotion by GrB of perforin-dependent and perforin-independent (anoikis) apoptosis, inflammation derived by the activation of some cytokines belonging to the IL-1 cytokine family, ECM remodeling, epithelial-to-mesenchymal transition (EMT) and fibrosis. A greater comprehension of the pathophysiological consequences of GrB-mediated multiple activities may favor the design of new therapies aim to inhibit different inflammatory pathological conditions such as inflammaging and age-related diseases, EMT and organ fibrosis.


Assuntos
Granzimas/imunologia , Inflamação/imunologia , Animais , Apoptose , Transição Epitelial-Mesenquimal , Matriz Extracelular , Fibrose , Humanos , Perforina/imunologia
19.
Exp Cell Res ; 396(1): 112260, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32890458

RESUMO

CD8+ T cells are considered a critical component of antitumor immunity. However, tumor-infiltrating CD8+ T cells may express more than one checkpoint molecules that have the potential to inhibit effector responses alone or cooperatively. Here, we focused on the expression dynamic of TIGIT and PD-1 in CD8+ T cells. TIGIT+ subset presented significantly higher PD-1 expression than TIGIT- subset in circulating CD8+ T cells. The expression dynamic of TIGIT and PD-1 was then tracked. In total CD8+ T cells, TIGIT mRNA increased more rapidly than PD-1 mRNA, and TIGIT+ CD8+ T cells upregulated PD-1 more rapidly than TIGIT- CD8+ T cells. Next, 24-h-stimulated CD8+ T cells were re-sorted into TIGIT+ and TIGIT- subsets, and the TIGIT+ cells that came from TIGIT- cells also presented significantly more rapid PD-1 induction than persistent TIGIT- CD8+ T cells. In non-small cell lung cancer (NSCLC) patients, the expression of PD-1 was more enriched in TIGIT+ cells than in TIGIT- cells in both circulating CD8+ T cells and tumor-infiltrating CD8+ T cells. Function analysis revealed that TIGIT+ CD8 T cells presented lower interferon-gamma, perforin 1, and granzyme B upregulation than TIGIT- CD8 T cells, especially in NSCLC patients. Overall, these data indicated that TIGIT presented earlier expression dynamic than PD-1 in activated CD8+ T cells and was upregulated in NSCLC patients.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Carcinoma Pulmonar de Células não Pequenas/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Receptor de Morte Celular Programada 1/genética , Receptores Imunológicos/genética , Linfócitos T CD8-Positivos/patologia , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Estudos de Casos e Controles , Feminino , Granzimas/genética , Granzimas/imunologia , Humanos , Imunofenotipagem , Interferon gama/genética , Interferon gama/imunologia , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Perforina/genética , Perforina/imunologia , Cultura Primária de Células , Receptor de Morte Celular Programada 1/imunologia , Receptores Imunológicos/imunologia , Transdução de Sinais
20.
J Immunol ; 205(9): 2519-2531, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32948683

RESUMO

Increasing evidence indicates that local hypofractionated radiotherapy (LRT) can elicit both immunogenic and immunosuppressive local and systemic immune responses. We thus hypothesized that blockade of LRT-induced immunosuppressive responses could augment the antitumor effects and induce an abscopal response. In this study, we found that the upregulation of Foxp3+ regulatory T cells (Tregs) in the mesothelioma tumor microenvironment after nonablative oligofractionated irradiation significantly limited the success of irradiation. Using DEREG mice, which allow conditional and efficient depletion of Foxp3+ Tregs by diphtheria toxin injection, we observed that transient Foxp3+ Treg depletion immediately after nonablative oligofractionated irradiation provided synergistic local control and biased the T cell repertoire toward central and effector memory T cells, resulting in long-term cure. Furthermore, this combination therapy showed significant abscopal effect on the nonirradiated tumors in a concomitant model of mesothelioma through systemic activation of cytotoxic T cells and enhanced production of IFN-γ and granzyme B. Although local control was preserved with one fraction of nonablative irradiation, three fractions were required to generate the abscopal effect. PD-1 and CTLA-4 were upregulated on tumor-infiltrating CD4+ and CD8+ T cells in irradiated and nonirradiated tumors, suggesting that immune checkpoint inhibitors could be beneficial after LRT and Foxp3+ Treg depletion. Our findings are applicable to the strategy of immuno-radiotherapy for generating optimal antitumor immune responses in the clinical setting. Targeting Tregs immediately after a short course of irradiation could have a major impact on the local response to irradiation and its abscopal effect.


Assuntos
Fatores de Transcrição Forkhead/imunologia , Mesotelioma Maligno/imunologia , Linfócitos T Reguladores/imunologia , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Antígeno CTLA-4/imunologia , Granzimas/imunologia , Imunidade/imunologia , Interferon gama/imunologia , Depleção Linfocítica/métodos , Linfócitos do Interstício Tumoral/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Receptor de Morte Celular Programada 1/imunologia , Linfócitos T Citotóxicos/imunologia , Microambiente Tumoral/imunologia
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