RESUMO
Hematopoiesis - the process by which blood cells are formed - has been studied intensely for over a century using a variety of model systems. There is conservation of the overall hematopoietic process between vertebrates, although some differences do exist. Over the last decade, the zebrafish has come to the forefront as a new model in hematopoiesis research, as it allows the use of large-scale genetics, chemical screens and transgenics. This comparative approach to understanding hematopoiesis has led to fundamental knowledge about the process and to the development of new therapies for disease. Here, we provide a broad overview of vertebrate hematopoiesis. We also highlight the benefits of using zebrafish as a model.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Hematopoese , Células-Tronco Hematopoéticas/citologia , Peixe-Zebra/sangue , Animais , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Desenvolvimento Embrionário , Fator de Transcrição GATA1/genética , Fator de Transcrição GATA1/metabolismo , Hemangioblastos/citologia , Hemangioblastos/metabolismo , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/metabolismo , Humanos , Prostaglandinas/metabolismo , Via de Sinalização Wnt , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismoRESUMO
Patients with type 2 diabetes (T2D) have disease-associated changes in B-cell function, but the role these changes play in disease pathogenesis is not well established. Data herein show B cells from obese mice produce a proinflammatory cytokine profile compared with B cells from lean mice. Complementary in vivo studies show that obese B cell-null mice have decreased systemic inflammation, inflammatory B- and T-cell cytokines, adipose tissue inflammation, and insulin resistance (IR) compared with obese WT mice. Reduced inflammation in obese/insulin resistant B cell-null mice associates with an increased percentage of anti-inflammatory regulatory T cells (Tregs). This increase contrasts with the sharply decreased percentage of Tregs in obese compared with lean WT mice and suggests that B cells may be critical regulators of T-cell functions previously shown to play important roles in IR. We demonstrate that B cells from T2D (but not non-T2D) subjects support proinflammatory T-cell function in obesity/T2D through contact-dependent mechanisms. In contrast, human monocytes increase proinflammatory T-cell cytokines in both T2D and non-T2D analyses. These data support the conclusion that B cells are critical regulators of inflammation in T2D due to their direct ability to promote proinflammatory T-cell function and secrete a proinflammatory cytokine profile. Thus, B cells are potential therapeutic targets for T2D.
Assuntos
Linfócitos B/imunologia , Citocinas/imunologia , Diabetes Mellitus Tipo 2/imunologia , Obesidade/imunologia , Linfócitos T Reguladores/imunologia , Animais , Linfócitos B/patologia , Diabetes Mellitus Tipo 2/patologia , Diabetes Mellitus Tipo 2/terapia , Feminino , Humanos , Inflamação/imunologia , Inflamação/patologia , Inflamação/terapia , Masculino , Camundongos , Camundongos Obesos , Obesidade/patologia , Obesidade/terapia , Linfócitos T Reguladores/patologiaRESUMO
Obesity and Type 2 diabetes mellitus (T2D) are characterized by pro-inflammatory alterations in the immune system including shifts in leukocyte subset differentiation and in cytokine/chemokine balance. The chronic, low-grade inflammation resulting largely from changes in T-cell, B-cell, and myeloid compartments promotes and/or exacerbates insulin resistance (IR) that, together with pancreatic islet failure, defines T2D. Animal model studies show that interruption of immune cell-mediated inflammation by any one of several methods almost invariably results in the prevention or delay of obesity and/or IR. However, anti-inflammatory therapies have had a modest impact on established T2D in clinical trials. These seemingly contradictory results indicate that a more comprehensive understanding of human IR/T2D-associated immune cell function is needed to leverage animal studies into clinical treatments. Important outstanding analyses include identifying potential immunological checkpoints in disease etiology, detailing immune cell/adipose tissue cross-talk, and defining strengths/weaknesses of model organism studies to determine whether we can harness the promising new field of immunometabolism to curb the global obesity and T2D epidemics.
Assuntos
Diabetes Mellitus Tipo 2/imunologia , Diabetes Mellitus Tipo 2/metabolismo , Metabolismo Energético , Obesidade/metabolismo , Adipócitos/imunologia , Tecido Adiposo/imunologia , Tecido Adiposo/metabolismo , Animais , Linfócitos B/imunologia , Diferenciação Celular , Humanos , Inflamação/imunologia , Resistência à Insulina , Leucócitos/imunologia , Camundongos , Obesidade/imunologia , Linfócitos T/imunologiaRESUMO
Mutational activation of KRAS occurs commonly in lung carcinogenesis and, with the recent FDA approval of covalent inhibitors of KRAS G12C such as sotorasib or adagrasib, KRAS oncoproteins are important pharmacological targets in non-small cell lung cancer (NSCLC). However, not all KRAS G12C -driven NSCLCs respond to these inhibitors, and the emergence of drug resistance in those patients that do respond can be rapid and pleiotropic. Hence, based on a backbone of covalent inhibition of KRAS G12C , efforts are underway to develop effective combination therapies. Here we report that inhibition of KRAS G12C signaling increases autophagy in KRAS G12C expressing lung cancer cells. Moreover, the combination of DCC-3116, a selective ULK1/2 inhibitor, plus sotorasib displays cooperative/synergistic suppression of human KRAS G12C -driven lung cancer cell proliferation in vitro and superior tumor control in vivo . Additionally, in genetically engineered mouse models of KRAS G12C -driven NSCLC, inhibition of either KRAS G12C or ULK1/2 decreases tumor burden and increases mouse survival. Consequently, these data suggest that ULK1/2-mediated autophagy is a pharmacologically actionable cytoprotective stress response to inhibition of KRAS G12C in lung cancer.
RESUMO
An immunosuppressive tumor microenvironment promotes tumor growth and is one of the main factors limiting the response to cancer immunotherapy. We have previously reported that inhibition of vacuolar protein sorting 34 (VPS34), a crucial lipid kinase in the autophagy/endosomal trafficking pathway, decreases tumor growth in several cancer models, increases infiltration of immune cells and sensitizes tumors to anti-programmed cell death protein 1/programmed cell death 1 ligand 1 therapy by upregulation of C-C motif chemokine 5 (CCL5) and C-X-C motif chemokine 10 (CXCL10) chemokines. The purpose of this study was to investigate the signaling mechanism leading to the VPS34-dependent chemokine increase. NanoString gene expression analysis was applied to tumors from mice treated with the VPS34 inhibitor SB02024 to identify key pathways involved in the anti-tumor response. We showed that VPS34 inhibitors increased the secretion of T-cell-recruitment chemokines in a cyclic GMP-AMP synthase (cGAS)/stimulator of interferon genes protein (STING)-dependent manner in cancer cells. Both pharmacological and small interfering RNA (siRNA)-mediated VPS34 inhibition increased cGAS/STING-mediated expression and secretion of CCL5 and CXCL10. The combination of VPS34 inhibitor and STING agonist further induced cytokine release in both human and murine cancer cells as well as monocytic or dendritic innate immune cells. Finally, the VPS34 inhibitor SB02024 sensitized B16-F10 tumor-bearing mice to STING agonist treatment and significantly improved mice survival. These results show that VPS34 inhibition augments the cGAS/STING pathway, leading to greater tumor control through immune-mediated mechanisms. We propose that pharmacological VPS34 inhibition may synergize with emerging therapies targeting the cGAS/STING pathway.
Assuntos
Classe III de Fosfatidilinositol 3-Quinases , Interferon Tipo I , Proteínas de Membrana , Transdução de Sinais , Animais , Proteínas de Membrana/metabolismo , Proteínas de Membrana/agonistas , Classe III de Fosfatidilinositol 3-Quinases/metabolismo , Humanos , Transdução de Sinais/efeitos dos fármacos , Camundongos , Linhagem Celular Tumoral , Interferon Tipo I/metabolismo , Nucleotidiltransferases/metabolismo , Camundongos Endogâmicos C57BL , Feminino , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêuticoRESUMO
An appropriate balance between proinflammatory (Th17 and Th1) and anti-inflammatory (regulatory T cells [Tregs] and Th2) subsets of T cells is critical to maintain homeostasis and avoid inflammatory disease. Type 2 diabetes (T2D) is a chronic inflammatory disease promoted by changes in immune cell function. Recent work indicates T cells are important mediators of inflammation in a mouse model of T2D. These studies identified an elevation in the Th17 and Th1 subsets with a decrease in the Treg subset, which culminates in inflammation and insulin resistance. Based on these data, we tested the hypothesis that T cells in T2D patients are skewed toward proinflammatory subsets. Our data show that blood from T2D patients has increased circulating Th17 cells and elevated activation of Th17 signature genes. Importantly, T cells required culture with monocytes to maintain Th17 signatures, and fresh ex vivo T cells from T2D patients appeared to be poised for IL-17 production. T cells from T2D patients also have increased production of IFN-γ, but produce healthy levels of IL-4. In contrast, T2D patients had decreased percentages of CD4(+) Tregs. These data indicate that T cells in T2D patients are naturally skewed toward proinflammatory subsets that likely promote chronic inflammation in T2D through elevated cytokine production. Potential therapies targeted toward resetting this balance need to be approached with caution due to the reciprocal relationship between Th17 cells and Tregs. Understanding the unique aspects of T2D T cells is essential to predict outcomes of such treatments.
Assuntos
Citocinas/biossíntese , Diabetes Mellitus Tipo 2/imunologia , Diabetes Mellitus Tipo 2/patologia , Mediadores da Inflamação/metabolismo , Monócitos/imunologia , Monócitos/patologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/patologia , Adulto , Idoso , Diferenciação Celular/imunologia , Células Cultivadas , Técnicas de Cocultura , Estudos Transversais , Diabetes Mellitus Tipo 2/metabolismo , Feminino , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/patologia , Interferon gama/biossíntese , Interleucina-17/biossíntese , Interleucina-17/metabolismo , Masculino , Pessoa de Meia-Idade , Monócitos/metabolismo , Subpopulações de Linfócitos T/metabolismo , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Auxiliares-Indutores/metabolismo , Linfócitos T Auxiliares-Indutores/patologiaRESUMO
OBJECTIVE: T cell inflammation plays pivotal roles in obesity-associated type 2 diabetes (T2DM). The identification of dominant sources of T cell inflammation in humans remains a significant gap in understanding disease pathogenesis. It was hypothesized that cytokine profiles from circulating T cells identify T cell subsets and T cell cytokines that define T2DM-associated inflammation. METHODS: Multiplex analyses were used to quantify T cell-associated cytokines in αCD3/αCD28-stimulated PBMCs, or B cell-depleted PBMCs, from subjects with T2DM or BMI-matched controls. Cytokine measurements were subjected to multivariate (principal component and partial least squares) analyses. Flow cytometry detected intracellular TNFα in multiple immune cell subsets in the presence/absence of antibodies that neutralize T cell cytokines. RESULTS: T cell cytokines were generally higher in T2DM samples, but Th17 cytokines are specifically important for classifying individuals correctly as T2DM. Multivariate analyses indicated that B cells support Th17 inflammation in T2DM but not control samples, while monocytes supported Th17 inflammation regardless of T2DM status. Partial least squares regression analysis indicated that both Th17 and Th1 cytokines impact %HbA1c. CONCLUSIONS: Among various T cell subsets, Th17 cells are major contributors to inflammation and hyperglycemia and are uniquely supported by B cells in obesity-associated T2DM.
Assuntos
Citocinas/imunologia , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/imunologia , Obesidade/complicações , Obesidade/imunologia , Células Th17/imunologia , Fator de Necrose Tumoral alfa/imunologia , Adulto , Idoso , Linfócitos B/imunologia , Células Cultivadas , Feminino , Humanos , Inflamação/complicações , Inflamação/imunologia , Leucócitos Mononucleares/imunologia , Masculino , Pessoa de Meia-Idade , Monócitos/imunologia , Adulto JovemRESUMO
Toll-like receptor 4 (TLR4) is an innate immune receptor that is constitutively and inducibly activated in monocytes. Although TLR4 is expressed at very low levels on human B cells from healthy individuals, recent reports showed that TLR4 expression and function is elevated in B cells from inflammatory disease patients. New data showed that TLR4 expression on B cells is increased upon stimulation through surface Igµ and CD40 in combination with IL-4. In contrast, monocyte stimulation through CD40 and IL-4 receptors decreased TLR4 surface expression. Analysis of molecular signatures of TLR4 activation in stimulated B cells suggested that TLR4 is regulated by different mechanisms in B cells compared to monocytes. PU.1 and interferon regulatory factor association with the TLR4 promoter are sufficient for TLR4 transcription, but are not sufficient for surface TLR4 expression on B cells. In contrast, the PU.1/IRF combination is sufficient for surface TLR4 expression on monocytes. These data identify mechanisms that can activate B cell TLR4 expression in inflammatory disease patients, and demonstrate that B cells have additional layers of TLR4 regulation absent in monocytes.