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1.
Methods Cell Biol ; 173: 33-48, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36653084

RESUMO

Natural killer (NK) cells are part of the innate immune system, the classic cytotoxic population of innate lymphoid cells (ILCs). They can directly kill virus-infected or tumor cells through different mechanisms without prior sensitization using their lytic functions in response to different signals (target cell ligands and/or inflammatory cytokines) and secreting cytokines, such as interferon gamma (IFNγ) and tumor necrosis factor (TNF). NK cells use antibody-dependent cell-mediated cytotoxicity (ADCC) to recognize and kill cells expressing target antigens when they are antibody coated. Redirected cytotoxicity is a technique used to target cells that do not per se activate NK cells. Here, we use redirected degranulation, a surrogate technique that correlates with redirected lysis. The P815 cell line (mouse mastocytoma) express fragment crystallizable gamma receptor II (FcγRII) and therefore could bind the Fc portion of mouse IgG antibodies, which through their fragment antigen-binding (Fab) may recognize NK cells activating receptors leading to target cell lysis. This technique could be used to determine the inhibitory or activating capacity of different receptors or isoforms and in immunotherapy using T cell and NK cell activators.


Assuntos
Imunidade Inata , Células Matadoras Naturais , Humanos , Camundongos , Animais , Células Matadoras Naturais/metabolismo , Citocinas/metabolismo , Citotoxicidade Celular Dependente de Anticorpos , Linhagem Celular
2.
Front Immunol ; 14: 1211839, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37575238

RESUMO

Natural killer (NK) cells are lymphocytes with potent antitumor functions and, consequently, several NK cell-based strategies have been developed for cancer immunotherapy. A remarkable therapeutic approach is the adoptive transfer of NK cells stimulated with IL-12, IL-15 and IL-18. This cytokine stimulation endows NK cells with properties that resemble immunological memory and, for this reason, they are known as cytokine-induced memory-like (CIML) NK cells. Very promising results have been reported in clinical trials and yet, there are still unknown aspects of CIML NK cells. Here, we have conducted a preliminary study of their mitochondrial dynamics. Our results show that upon IL-12/15/18 stimulation the viability of NK cells decreased and an increment in mitochondrial superoxide levels was observed. In addition, we found that mitochondria appeared slightly elongated and their cristae density decreased following IL-12/15/18 stimulation, possibly in a process mediated by the low levels of optic atrophy type 1 (OPA1) protein. Interestingly, although mitophagy was slightly impaired, an increase in autophagic flux was observed, which might explain the reduced viability and the accumulation of unfit mitochondria. Our findings could be of relevance in order to design new strategies intended to improve the mitochondrial fitness of IL-12/15/18-stimulated NK cells with the aim of improving their therapeutic efficacy.


Assuntos
Interleucina-18 , Dinâmica Mitocondrial , Humanos , Interleucina-18/farmacologia , Citocinas/metabolismo , Células Matadoras Naturais , Interleucina-12
3.
Front Immunol ; 13: 884648, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35603208

RESUMO

Natural killer (NK) cells are lymphocytes with a key role in the defense against viral infections and tumor cells. Although NK cells are classified as innate lymphoid cells (ILCs), under certain circumstances they exhibit adaptive and memory-like features. The latter may be achieved, among others, by a brief stimulation with interleukin (IL)-12, IL-15 and IL-18. These cytokine-induced memory-like (CIML) NK cells resemble the trained immunity observed in myeloid cells. CIML NK cells undergo transcriptional, epigenetic and metabolic reprogramming that, along with changes in the expression of cell surface receptors and components of cytotoxic granules, are responsible for their enhanced effector functions after a resting period. In addition, these memory-like NK cells persist for a long time, which make them a good candidate for cancer immunotherapy. Currently, several clinical trials are testing CIML NK cells infusions to treat tumors, mostly hematological malignancies. In relapse/refractory acute myeloid leukemia (AML), the adoptive transfer of CIML NK cells is safe and complete clinical remissions have been observed. In our review, we sought to summarize the current knowledge about the generation and molecular basis of NK cell memory-like responses and the up-to-date results from clinical trials with CIML NK cells.


Assuntos
Citocinas , Imunidade Inata , Citocinas/metabolismo , Memória Imunológica , Interleucina-12/farmacologia , Células Matadoras Naturais
4.
iScience ; 25(10): 105235, 2022 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-36262311

RESUMO

Autologous hematopoietic stem cell transplantation (autoHSCT) is a treatment option for hematological disorders and pediatric solid tumors. After an autoHSCT, natural killer (NK) cells are the first lymphocyte subset returning to normal levels. To uncover global changes during NK cell reconstitution after autoHSCT, we performed RNA-sequencing on NK cells before and after autoHSCT. Results showed profound changes in the gene expression profile of NK cells immediately after autoHSCT. Several biological processes including cell cycle, DNA replication and the mevalonate pathway were enriched. Significantly, we observed that following autoHSCT, NK cells acquired a decidual-like gene expression profile, including the expression of CD9. By using multiparametric flow cytometry, we confirmed the expansion of NK cells expressing CD9 immediately after autoHSCT, which exhibited higher granzyme B and perforin expression levels than CD9- NK cells. These results provide insights into the physiopathology of NK cells during their reconstitution after autoHSCT.

5.
Cancers (Basel) ; 13(7)2021 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-33808201

RESUMO

Natural killer (NK) cells are phenotypically and functionally diverse lymphocytes with the ability to recognize and kill malignant cells without prior sensitization, and therefore, they have a relevant role in tumor immunosurveillance. NK cells constitute the main lymphocyte subset in peripheral blood in the first week after hematopoietic stem cell transplantation (HSCT). Although the role that NK cells play in allogenic HSCT settings has been documented for years, their significance and beneficial effects associated with the outcome after autologous HSCT are less recognized. In this review, we have summarized fundamental aspects of NK cell biology, such as, NK cell subset diversity, their effector functions, and differentiation. Moreover, we have reviewed the factors that affect autologous HSCT outcome, with particular attention to the role played by NK cells and their receptor repertoire in this regard.

6.
Front Pediatr ; 9: 734645, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34912758

RESUMO

Background: Infants with severe atopic dermatitis (AD) may be sensitized to foods that have not been introduced into their diet, posing a risk for developing an immediate hypersensitivity reaction on the first exposure to the food to which they are sensitized. The aim of this work was to perform an analysis of the sensitization profile in infants with moderate-to-severe AD and to identify cellular and molecular markers for food allergy (FA). Methods: Blood samples from healthy donors and children with moderate-to-severe AD were studied. Specific IgE to several allergens were determined using ImmunoCAP FEIA system and ISAC technology. Furthermore, using flow cytometry-based studies, basophils and regulatory T (Treg) cells were phenotypically characterized. Results: 90% of children with AD were sensitized to food antigens before introducing them into the diet, and 100% developed FA. Phenotypic analysis showed a significantly higher percentage of CTLA-4 and PD-1 expressing Treg cells in AD patients than in healthy controls. Basophils from patients exhibited a marked reduction in the expression of CD300a, higher expression of FcεRI and CXCR4, and to some extent higher expression of CD63 and CD300c. Conclusions: Infants with moderate-to-severe AD are at high risk of being sensitized to food allergens. Therefore, to avoid allergic reactions, broad-spectrum sensitization studies are necessary before introducing complementary diet. Increased expression of CTLA-4 and PD-1 suggests greater suppressive potential of Treg cells in infants with AD than healthy controls. Furthermore, our results suggest a role for CD300 molecules on circulating basophils as possible biomarkers for FA susceptibility.

7.
Front Immunol ; 12: 748207, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34675932

RESUMO

Autologous hematopoietic stem cell transplantation (autoHSCT) is a standard of care for transplant-eligible patients with multiple myeloma (MM). Among factors that influence outcome after autoHSCT, it has been suggested that the number of natural killer (NK) cells plays an important role. However, the impact that different NK cell subsets and their phenotype could have in disease progression after autoHSCT are less clear. For this reason, we have phenotypically and functionally characterized NK cells during immune system reconstitution after autoHSCT in 54 MM patients. Shortly after leukocyte recovery, an extensive redistribution of NK cell subsets occurs in these patients. In addition, NK cells undergo a profound phenotypic change characterized, among others, by their increased proliferative capacity and immature phenotype. Importantly, MM patients who showed lower frequencies of the mature highly differentiated NKG2A-CD57+ NK cell subset at +30 and +100 days after autoHSCT experienced superior progression-free survival and had a longer time to the next treatment than those with higher frequencies. Our results provide significant insights into NK cell reconstitution after autoHSCT and suggest that the degree of NK cell maturation after autoHSCT affects the clinical outcome of MM patients treated with this therapeutic strategy.


Assuntos
Transplante de Células-Tronco Hematopoéticas , Células Matadoras Naturais/citologia , Mieloma Múltiplo/imunologia , Adulto , Idoso , Citotoxicidade Imunológica , Feminino , Humanos , Interleucina-15/sangue , Estimativa de Kaplan-Meier , Células Matadoras Naturais/imunologia , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/sangue , Mieloma Múltiplo/mortalidade , Mieloma Múltiplo/terapia , Modelos de Riscos Proporcionais , Transplante Autólogo , Resultado do Tratamento
8.
Front Immunol ; 12: 655934, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33777054

RESUMO

COVID-19 manifests with a wide diversity of clinical phenotypes characterized by dysfunctional and exaggerated host immune responses. Many results have been described on the status of the immune system of patients infected with SARS-CoV-2, but there are still aspects that have not been fully characterized or understood. In this study, we have analyzed a cohort of patients with mild, moderate and severe disease. We performed flow cytometric studies and correlated the data with the clinical characteristics and clinical laboratory values of the patients. Both conventional and unsupervised data analyses concluded that patients with severe disease are characterized, among others, by a higher state of activation in all T cell subsets (CD4, CD8, double negative and T follicular helper cells), higher expression of perforin and granzyme B in cytotoxic cells, expansion of adaptive NK cells and the accumulation of activated and immature dysfunctional monocytes which are identified by a low expression of HLA-DR and an intriguing shift in the expression pattern of CD300 receptors. More importantly, correlation analysis showed a strong association between the alterations in the immune cells and the clinical signs of severity. These results indicate that patients with severe COVID-19 have a broad perturbation of their immune system, and they will help to understand the immunopathogenesis of COVID-19.


Assuntos
COVID-19/imunologia , Citotoxicidade Imunológica , Células Matadoras Naturais/imunologia , Ativação Linfocitária , Monócitos/imunologia , Receptores Imunológicos/sangue , SARS-CoV-2/imunologia , Linfócitos T/imunologia , Idoso , COVID-19/sangue , COVID-19/diagnóstico , COVID-19/virologia , Estudos de Casos e Controles , Estudos Transversais , Feminino , Citometria de Fluxo , Interações Hospedeiro-Patógeno , Humanos , Imunofenotipagem , Células Matadoras Naturais/metabolismo , Células Matadoras Naturais/virologia , Masculino , Pessoa de Meia-Idade , Monócitos/metabolismo , Monócitos/virologia , Fenótipo , Índice de Gravidade de Doença , Linfócitos T/metabolismo , Linfócitos T/virologia
9.
STAR Protoc ; 1(3): 100149, 2020 12 18.
Artigo em Inglês | MEDLINE | ID: mdl-33377043

RESUMO

Although scarce in the peripheral blood of healthy people, CD56neg NK cells are known to be expanded in some pathological conditions. However, studies on CD56neg NK cells had revealed contradictions, probably due to the lack of a specific NK cell surface marker that helps to identify this subset. This protocol details the step-by-step procedure for the identification and functional analysis of CD56neg NK cells, providing an improved gating strategy for the selection of this intriguing population. For complete details on the use and execution of this protocol, please refer to Orrantia et al. (2020).


Assuntos
Antígeno CD56/metabolismo , Citometria de Fluxo/métodos , Células Matadoras Naturais/citologia , Bioensaio , Humanos , Células K562 , Coloração e Rotulagem
10.
Semin Hematol ; 57(4): 213-224, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-33256914

RESUMO

Natural killer (NK) cells are lymphocytes with potent antitumor functions and, therefore, multiple NK cell-based cancer immunotherapies have been developed and are currently being tested. However, there is a necessity to find new means to improve these therapies, and immunometabolism represents an attractive target. NK cell effector functions are intricately linked to their metabolism, and modulating the latter could be the key to release their full potential. In this review, we have summarized how NK cell metabolism is regulated during some processes, such as maturation, viral infection, and cytokine stimulation. Additionally, we provide an overview of how NK cell metabolism is affected by current therapeutic approaches aimed to promote NK cell expansion and/or to increase their effector functions. We have also recapitulated several strategies that could help alleviating the metabolic impairment that characterizes tumor-infiltrating NK cells, and thus increase or restore their effector functions. Furthermore, we have reviewed several therapeutic approaches targeting cancer metabolism that could synergize with NK cell-based cancer immunotherapies, and thus enhance their efficacy.


Assuntos
Imunoterapia/métodos , Células Matadoras Naturais/metabolismo , Neoplasias/terapia , Humanos , Neoplasias/imunologia
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