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2.
J Transl Autoimmun ; 6: 100185, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36654851

RESUMO

Dual-specificity tyrosine phosphorylation-regulated kinase 1 A (DYRK1A) has been proposed as a novel regulator of adaptive immune homeostasis through modulating T cell polarization. Thus, DYRK1A could present a potential target in autoimmune disorders. Here, we identify FRTX-02 as a novel compound exhibiting potent and selective inhibition of DYRK1A. FRTX-02 induced transcriptional activity of the DYRK1A substrate NFAT in T cell lines. Correspondingly, FRTX-02 promoted ex vivo CD4+ polarization into anti-inflammatory Tregs and reduced their polarization into pro-inflammatory Th1 or Th17 cells. We show that FRTX-02 could also limit innate immune responses through negative regulation of the MyD88/IRAK4-NF-κB axis in a mast cell line. Finally, in mouse models of psoriasis and atopic dermatitis, both oral and topical formulations of FRTX-02 reduced inflammation and disease biomarkers in a dose-dependent manner. These results support further studies of DYRK1A inhibitors, including FRTX-02, as potential therapies for chronic inflammatory and autoimmune conditions.

3.
Sci Transl Med ; 14(627): eabi4888, 2022 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-35020411

RESUMO

Individuals with Down syndrome show cellular and clinical features of dysregulated aging of the immune system, including a shift from naïve to memory T cells and increased incidence of autoimmunity. However, a quantitative understanding of how various immune compartments change with age in Down syndrome remains lacking. Here, we performed deep immunophenotyping of a cohort of individuals with Down syndrome across the life span, selecting for autoimmunity-free individuals. We simultaneously interrogated age- and sex-matched healthy controls and people with type 1 diabetes as a representative autoimmune disease. We built an analytical software, IMPACD (Iterative Machine-assisted Permutational Analysis of Cytometry Data), that enabled us to rapidly identify many features of immune dysregulation in Down syndrome shared with other autoimmune diseases. We found quantitative and qualitative dysregulation of naïve CD4+ and CD8+ T cells in individuals with Down syndrome and identified interleukin-6 as a candidate driver of some of these changes, thus extending the consideration of immunopathologic cytokines in Down syndrome beyond interferons. We used immune cellular composition to generate three linear models of aging (immune clocks) trained on control participants. All three immune clocks demonstrated advanced immune aging in individuals with Down syndrome. One of these clocks, informed by Down syndrome­relevant biology, also showed advanced immune aging in individuals with type 1 diabetes. Orthologous RNA sequencing­derived immune clocks also demonstrated advanced immune aging in individuals with Down syndrome. Together, our findings demonstrate an approach to studying immune aging in Down syndrome that may have implications in other autoimmune diseases.


Assuntos
Doenças Autoimunes , Diabetes Mellitus Tipo 1 , Síndrome de Down , Envelhecimento , Autoimunidade/genética , Linfócitos T CD8-Positivos , Síndrome de Down/genética , Humanos , Imunofenotipagem
4.
Elife ; 102021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34378531

RESUMO

Maintenance of immune homeostasis involves a synergistic relationship between the host and the microbiome. Canonical interferon (IFN) signaling controls responses to acute microbial infection, through engagement of the STAT1 transcription factor. However, the contribution of tonic levels of IFN to immune homeostasis in the absence of acute infection remains largely unexplored. We report that STAT1 KO mice spontaneously developed an inflammatory disease marked by myeloid hyperplasia and splenic accumulation of hematopoietic stem cells. Moreover, these animals developed inflammatory bowel disease. Profiling gut bacteria revealed a profound dysbiosis in the absence of tonic IFN signaling, which triggered expansion of TH17 cells and loss of splenic Treg cells. Reduction of bacterial load by antibiotic treatment averted the TH17 bias and blocking IL17 signaling prevented myeloid expansion and splenic stem cell accumulation. Thus, tonic IFNs regulate gut microbial ecology, which is crucial for maintaining physiologic immune homeostasis and preventing inflammation.


Assuntos
Disbiose/imunologia , Microbioma Gastrointestinal , Inflamação/genética , Interferons/administração & dosagem , Interleucina-17/genética , Fator de Transcrição STAT1/genética , Animais , Feminino , Interleucina-17/metabolismo , Camundongos , Camundongos Knockout , Fator de Transcrição STAT1/metabolismo
5.
Transl Sci Rare Dis ; 5(3-4): 99-129, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34268067

RESUMO

BACKGROUND: Recent advances in medical care have increased life expectancy and improved the quality of life for people with Down syndrome (DS). These advances are the result of both pre-clinical and clinical research but much about DS is still poorly understood. In 2020, the NIH announced their plan to update their DS research plan and requested input from the scientific and advocacy community. OBJECTIVE: The National Down Syndrome Society (NDSS) and the LuMind IDSC Foundation worked together with scientific and medical experts to develop recommendations for the NIH research plan. METHODS: NDSS and LuMind IDSC assembled over 50 experts across multiple disciplines and organized them in eleven working groups focused on specific issues for people with DS. RESULTS: This review article summarizes the research gaps and recommendations that have the potential to improve the health and quality of life for people with DS within the next decade. CONCLUSIONS: This review highlights many of the scientific gaps that exist in DS research. Based on these gaps, a multidisciplinary group of DS experts has made recommendations to advance DS research. This paper may also aid policymakers and the DS community to build a comprehensive national DS research strategy.

6.
Mol Cell Biol ; 41(9): e0008521, 2021 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-34124936

RESUMO

Immune health requires innate and adaptive immune cells to engage precisely balanced pro- and anti-inflammatory forces. We employ the concept of chemical immunophenotypes to classify small molecules functionally or mechanistically according to their patterns of effects on primary innate and adaptive immune cells. The high-specificity, low-toxicity cyclin-dependent kinase 8 (CDK8) inhibitor 16-didehydro-cortistatin A (DCA) exerts a distinct tolerogenic profile in both innate and adaptive immune cells. DCA promotes regulatory T cells (Treg) and Th2 differentiation while inhibiting Th1 and Th17 differentiation in both murine and human cells. This unique chemical immunophenotype led to mechanistic studies showing that DCA promotes Treg differentiation in part by regulating a previously undescribed CDK8-GATA3-FOXP3 pathway that regulates early pathways of Foxp3 expression. These results highlight previously unappreciated links between Treg and Th2 differentiation and extend our understanding of the transcription factors that regulate Treg differentiation and their temporal sequencing. These findings have significant implications for future mechanistic and translational studies of CDK8 and CDK8 inhibitors.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Quinase 8 Dependente de Ciclina/antagonistas & inibidores , Fatores de Transcrição Forkhead/metabolismo , Fator de Transcrição GATA3/metabolismo , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Tolerância Imunológica/efeitos dos fármacos , Imunofenotipagem , Isoquinolinas/farmacologia , Adolescente , Adulto , Animais , Linfócitos T CD4-Positivos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Quinase 8 Dependente de Ciclina/metabolismo , Humanos , Imunidade Inata/efeitos dos fármacos , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-jun/metabolismo , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais/efeitos dos fármacos , Adulto Jovem
8.
J Immunol ; 205(2): 414-424, 2020 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-32522834

RESUMO

Genome-wide association studies have identified common genetic variants impacting human diseases; however, there are indications that the functional consequences of genetic polymorphisms can be distinct depending on cell type-specific contexts, which produce divergent phenotypic outcomes. Thus, the functional impact of genetic variation and the underlying mechanisms of disease risk are modified by cell type-specific effects of genotype on pathological phenotypes. In this study, we extend these concepts to interrogate the interdependence of cell type- and stimulation-specific programs influenced by the core autophagy gene Atg16L1 and its T300A coding polymorphism identified by genome-wide association studies as linked with increased risk of Crohn's disease. We applied a stimulation-based perturbational profiling approach to define Atg16L1 T300A phenotypes in dendritic cells and T lymphocytes. Accordingly, we identified stimulus-specific transcriptional signatures revealing T300A-dependent functional phenotypes that mechanistically link inflammatory cytokines, IFN response genes, steroid biosynthesis, and lipid metabolism in dendritic cells and iron homeostasis and lysosomal biogenesis in T lymphocytes. Collectively, these studies highlight the combined effects of Atg16L1 genetic variation and stimulatory context on immune function.


Assuntos
Proteínas Relacionadas à Autofagia/metabolismo , Doença de Crohn/metabolismo , Células Dendríticas/fisiologia , Genótipo , Linfócitos T/fisiologia , Animais , Proteínas Relacionadas à Autofagia/genética , Células Cultivadas , Doença de Crohn/genética , Predisposição Genética para Doença , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Especificidade de Órgãos , Fenótipo , Polimorfismo Genético , Risco , Ativação Transcricional
9.
J Immunol ; 203(7): 1820-1829, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31451676

RESUMO

The clear role of autophagy in human inflammatory diseases such as Crohn disease was first identified by genome-wide association studies and subsequently dissected in multiple mechanistic studies. ATG16L1 has been particularly well studied in knockout and hypomorph settings as well as models recapitulating the Crohn disease-associated T300A polymorphism. Interestingly, ATG16L1 has a single homolog, ATG16L2, which is independently implicated in diseases, including Crohn disease and systemic lupus erythematosus. However, the contribution of ATG16L2 to canonical autophagy pathways and other cellular functions is poorly understood. To better understand its role, we generated and analyzed the first, to our knowledge, ATG16L2 knockout mouse. Our results show that ATG16L1 and ATG16L2 contribute very distinctly to autophagy and cellular ontogeny in myeloid, lymphoid, and epithelial lineages. Dysregulation of any of these lineages could contribute to complex diseases like Crohn disease and systemic lupus erythematosus, highlighting the value of examining cell-specific effects. We also identify a novel genetic interaction between ATG16L2 and epithelial ATG16L1. These findings are discussed in the context of how these genes may contribute distinctly to human disease.


Assuntos
Morte Celular Autofágica , Proteínas Relacionadas à Autofagia , Proteínas de Transporte , Doença de Crohn , Lúpus Eritematoso Sistêmico , Animais , Morte Celular Autofágica/genética , Morte Celular Autofágica/imunologia , Proteínas Relacionadas à Autofagia/genética , Proteínas Relacionadas à Autofagia/imunologia , Proteínas de Transporte/genética , Proteínas de Transporte/imunologia , Doença de Crohn/genética , Doença de Crohn/imunologia , Modelos Animais de Doenças , Humanos , Lúpus Eritematoso Sistêmico/genética , Lúpus Eritematoso Sistêmico/imunologia , Camundongos , Camundongos Knockout , Especificidade de Órgãos/genética , Especificidade de Órgãos/imunologia
10.
Nat Chem Biol ; 13(10): 1102-1108, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28805801

RESUMO

Enhancing production of the anti-inflammatory cytokine interleukin-10 (IL-10) is a promising strategy to suppress pathogenic inflammation. To identify new mechanisms regulating IL-10 production, we conducted a phenotypic screen for small molecules that enhance IL-10 secretion from activated dendritic cells. Mechanism-of-action studies using a prioritized hit from the screen, BRD6989, identified the Mediator-associated kinase CDK8, and its paralog CDK19, as negative regulators of IL-10 production during innate immune activation. The ability of BRD6989 to upregulate IL-10 is recapitulated by multiple, structurally differentiated CDK8 and CDK19 inhibitors and requires an intact cyclin C-CDK8 complex. Using a highly parallel pathway reporter assay, we identified a role for enhanced AP-1 activity in IL-10 potentiation following CDK8 and CDK19 inhibition, an effect associated with reduced phosphorylation of a negative regulatory site on c-Jun. These findings identify a function for CDK8 and CDK19 in regulating innate immune activation and suggest that these kinases may warrant consideration as therapeutic targets for inflammatory disorders.


Assuntos
Quinase 8 Dependente de Ciclina/metabolismo , Interleucina-10/biossíntese , Células Mieloides/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Células Cultivadas , Quinase 8 Dependente de Ciclina/imunologia , Relação Dose-Resposta a Droga , Humanos , Interleucina-10/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Células Mieloides/imunologia , Células Mieloides/metabolismo , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-Atividade
11.
Transfus Med Rev ; 31(1): 36-44, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27523957

RESUMO

The balanced differentiation of naive CD4+ T cells into either pro- or anti-inflammatory fates is a central regulator of immune homeostasis, dysregulation of which can lead to inflammatory disease or cancer. Accordingly, the development of diagnostics and therapeutics to measure and modulate this balance is of great interest. In this review, we focus on the predominant anti-inflammatory subset, regulatory T cells, discussing key concepts including development, function, antigen specificity, and lineage stability. In particular, we highlight how these notions are shaping the evolution of therapeutics, especially in the context of the transfusion medicine specialist, and identify several key areas that urgently need to be addressed.


Assuntos
Bioengenharia/métodos , Técnicas de Cultura de Células/métodos , Imunoterapia Adotiva/métodos , Linfócitos T Reguladores , Imunologia de Transplantes , Bioengenharia/tendências , Técnicas de Cultura de Células/tendências , Humanos , Imunoterapia Adotiva/tendências , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/fisiologia , Linfócitos T Reguladores/transplante , Medicina Transfusional/métodos , Medicina Transfusional/tendências , Reação Transfusional/sangue , Reação Transfusional/imunologia
12.
ACS Chem Biol ; 11(8): 2105-11, 2016 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-27224444

RESUMO

Salt-inducible kinases (SIKs) are promising therapeutic targets for modulating cytokine responses during innate immune activation. The study of SIK inhibition in animal models of disease has been limited by the lack of selective small-molecule probes suitable for modulating SIK function in vivo. We used the pan-SIK inhibitor HG-9-91-01 as a starting point to develop improved analogs, yielding a novel probe 5 (YKL-05-099) that displays increased selectivity for SIKs versus other kinases and enhanced pharmacokinetic properties. Well-tolerated doses of YKL-05-099 achieve free serum concentrations above its IC50 for SIK2 inhibition for >16 h and reduce phosphorylation of a known SIK substrate in vivo. While in vivo active doses of YKL-05-099 recapitulate the effects of SIK inhibition on inflammatory cytokine responses, they did not induce metabolic abnormalities observed in Sik2 knockout mice. These results identify YKL-05-099 as a useful probe to investigate SIK function in vivo and further support the development of SIK inhibitors for treatment of inflammatory disorders.


Assuntos
Sondas Moleculares/química , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Células Cultivadas , Concentração Inibidora 50 , Camundongos , Camundongos Knockout , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/química
13.
Am J Hematol ; 91(1): 46-9, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26492443

RESUMO

Factor V Leiden (FVLeiden ) is a common hereditary thrombophilia that causes activated protein C (APC) resistance. This review describes many of the most fascinating features of FVLeiden , including background features, mechanisms of hypercoagulability, the founder mutation concept, the "FVLeiden paradox," synergistic interaction with other thrombotic risk factors, the intertwined relationship between FVLeiden and APC resistance testing, and other, uncommon mutations implicated in causing APC resistance. In addition, there are several conditions where laboratory tests for APC resistance and FVLeiden are or can be discrepant, including lupus anticoagulants, anticoagulants such as direct thrombin inhibitors (dabigatran, argatroban, and bivalirudin) and rivaroxaban, as well as pseudohomozygous, pseudo-wildtype, liver transplant, and bone marrow transplant patients. The laboratory test error rate for FVLeiden is also presented.


Assuntos
Resistência à Proteína C Ativada , Fator V/genética , Resistência à Proteína C Ativada/sangue , Resistência à Proteína C Ativada/etiologia , Resistência à Proteína C Ativada/genética , Anticoagulantes/uso terapêutico , Coagulação Sanguínea , Testes de Coagulação Sanguínea , Análise Mutacional de DNA , Humanos , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/complicações , Mutação Puntual
14.
Elife ; 42015 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-25998054

RESUMO

The balance between Th17 and T regulatory (Treg) cells critically modulates immune homeostasis, with an inadequate Treg response contributing to inflammatory disease. Using an unbiased chemical biology approach, we identified a novel role for the dual specificity tyrosine-phosphorylation-regulated kinase DYRK1A in regulating this balance. Inhibition of DYRK1A enhances Treg differentiation and impairs Th17 differentiation without affecting known pathways of Treg/Th17 differentiation. Thus, DYRK1A represents a novel mechanistic node at the branch point between commitment to either Treg or Th17 lineages. Importantly, both Treg cells generated using the DYRK1A inhibitor harmine and direct administration of harmine itself potently attenuate inflammation in multiple experimental models of systemic autoimmunity and mucosal inflammation. Our results identify DYRK1A as a physiologically relevant regulator of Treg cell differentiation and suggest a broader role for other DYRK family members in immune homeostasis. These results are discussed in the context of human diseases associated with dysregulated DYRK activity.


Assuntos
Diferenciação Celular/imunologia , Homeostase/imunologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Tirosina Quinases/metabolismo , Linfócitos T Reguladores/metabolismo , Células Th17/metabolismo , Animais , Técnicas de Cultura de Células , Harmina/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/genética , Quinases Dyrk
15.
Am J Hematol ; 89(12): 1147-50, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25293789

RESUMO

Activated protein C resistance assays can detect factor V Leiden with high accuracy, depending on the method used. Factor Xa inhibitors such as rivaroxaban and direct thrombin inhibitors including dabigatran, argatroban, and bivalirudin can cause falsely normal results. Lupus anticoagulants can cause incorrect results in most current assays. Assays that include dilution into factor V-deficient plasma are needed to avoid interference from factor deficiencies or elevations, which can arise from a wide variety of conditions such as warfarin, liver dysfunction, or pregnancy. The pros and cons of the currently available assays are discussed.


Assuntos
Resistência à Proteína C Ativada/diagnóstico , Bioensaio/normas , Fator V/análise , Proteína C/metabolismo , Resistência à Proteína C Ativada/sangue , Adulto , Antitrombinas/química , Arginina/análogos & derivados , Benzimidazóis/química , Testes de Coagulação Sanguínea , Criança , Dabigatrana , Fator V/metabolismo , Fator Xa/metabolismo , Reações Falso-Positivas , Feminino , Hirudinas/química , Humanos , Inibidor de Coagulação do Lúpus/química , Morfolinas/química , Fragmentos de Peptídeos/química , Ácidos Pipecólicos/química , Gravidez , Proteínas Recombinantes/química , Rivaroxabana , Sulfonamidas , Tiofenos/química , Varfarina/química , beta-Alanina/análogos & derivados , beta-Alanina/química
16.
Proc Natl Acad Sci U S A ; 111(34): 12468-73, 2014 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-25114223

RESUMO

Genetic alterations that reduce the function of the immunoregulatory cytokine IL-10 contribute to colitis in mouse and man. Myeloid cells such as macrophages (MΦs) and dendritic cells (DCs) play an essential role in determining the relative abundance of IL-10 versus inflammatory cytokines in the gut. As such, using small molecules to boost IL-10 production by DCs-MΦs represents a promising approach to increase levels of this cytokine specifically in gut tissues. Toward this end, we screened a library of well-annotated kinase inhibitors for compounds that enhance production of IL-10 by murine bone-marrow-derived DCs stimulated with the yeast cell wall preparation zymosan. This approach identified a number of kinase inhibitors that robustly up-regulate IL-10 production including the Food and Drug Administration (FDA)-approved drugs dasatinib, bosutinib, and saracatinib that target ABL, SRC-family, and numerous other kinases. Correlating the kinase selectivity profiles of the active compounds with their effect on IL-10 production suggests that inhibition of salt-inducible kinases (SIKs) mediates the observed IL-10 increase. This was confirmed using the SIK-targeting inhibitor HG-9-91-01 and a series of structural analogs. The stimulatory effect of SIK inhibition on IL-10 is also associated with decreased production of the proinflammatory cytokines IL-1ß, IL-6, IL-12, and TNF-α, and these coordinated effects are observed in human DCs-MΦs and anti-inflammatory CD11c(+) CX3CR1(hi) cells isolated from murine gut tissue. Collectively, these studies demonstrate that SIK inhibition promotes an anti-inflammatory phenotype in activated myeloid cells marked by robust IL-10 production and establish these effects as a previously unidentified activity associated with several FDA-approved multikinase inhibitors.


Assuntos
Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Interleucina-10/biossíntese , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Compostos de Anilina/farmacologia , Animais , Células Cultivadas , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Citocinas/biossíntese , Dasatinibe , Células Dendríticas/enzimologia , Avaliação Pré-Clínica de Medicamentos , Humanos , Mediadores da Inflamação/metabolismo , Doenças Inflamatórias Intestinais/tratamento farmacológico , Doenças Inflamatórias Intestinais/enzimologia , Doenças Inflamatórias Intestinais/imunologia , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/enzimologia , Intestino Delgado/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células Mieloides/efeitos dos fármacos , Células Mieloides/enzimologia , Células Mieloides/imunologia , Nitrilas/farmacologia , Compostos de Fenilureia/química , Compostos de Fenilureia/farmacologia , Inibidores de Proteínas Quinases/química , Pirimidinas/química , Pirimidinas/farmacologia , Quinolinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/enzimologia , Linfócitos T Reguladores/imunologia , Tiazóis/farmacologia , Fatores de Transcrição/metabolismo
17.
ACS Chem Biol ; 8(12): 2724-2733, 2013 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-24168452

RESUMO

Autophagy is an evolutionarily conserved catabolic process that directs cytoplasmic proteins, organelles and microbes to lysosomes for degradation. Autophagy acts at the intersection of pathways involved in cellular stress, host defense, and modulation of inflammatory and immune responses; however, the details of how the autophagy network intersects with these processes remain largely undefined. Given the role of autophagy in several human diseases, it is important to determine the extent to which modulators of autophagy also modify inflammatory or immune pathways and whether it is possible to modulate a subset of these pathways selectively. Here, we identify small-molecule inducers of basal autophagy (including several FDA-approved drugs) and characterize their effects on IL-1ß production, autophagic engulfment and killing of intracellular bacteria, and development of Treg, TH17, and TH1 subsets from naïve T cells. Autophagy inducers with distinct, selective activity profiles were identified that reveal the functional architecture of connections between autophagy, and innate and adaptive immunity. In macrophages from mice bearing a conditional deletion of the essential autophagy gene Atg16L1, the small molecules inhibit IL-1ß production to varying degrees suggesting that individual compounds may possess both autophagy-dependent and autophagy-independent activity on immune pathways. The small molecule autophagy inducers constitute useful probes to test the contributions of autophagy-related pathways in diseases marked by impaired autophagy or elevated IL-1ß and to test novel therapeutic hypotheses.


Assuntos
Imunidade Adaptativa/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Imunidade Inata/efeitos dos fármacos , Fatores Imunológicos/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Autofagia/imunologia , Proteínas Relacionadas à Autofagia , Proteínas de Transporte/genética , Proteínas de Transporte/imunologia , Regulação da Expressão Gênica , Células HeLa , Ensaios de Triagem em Larga Escala , Humanos , Fatores Imunológicos/química , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Transdução de Sinais , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-Atividade , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Células Th1/citologia , Células Th1/efeitos dos fármacos , Células Th1/imunologia , Células Th17/citologia , Células Th17/efeitos dos fármacos , Células Th17/imunologia
18.
Autophagy ; 9(4): 528-37, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23327930

RESUMO

Autophagy is a conserved homeostatic process in which cytoplasmic contents are degraded and recycled. Two ubiquitin-like conjugation pathways are required for the generation of autophagosomes, and ATG5 is necessary for both of these processes. Studies of mice deficient in ATG5 reveal a key role for autophagy in T lymphocyte function, as well as in B cell development and B-1a B cell maintenance. However, the role of autophagy genes in B cell function and antibody production has not been described. Using mice in which Atg5 is conditionally deleted in B lymphocytes, we showed here that this autophagy gene is essential for plasma cell homeostasis. In the absence of B cell ATG5 expression, antibody responses were significantly diminished during antigen-specific immunization, parasitic infection and mucosal inflammation. Atg5-deficient B cells maintained the ability to produce immunoglobulin and undergo class-switch recombination, yet had impaired SDC1 expression, significantly decreased antibody secretion in response to toll-like receptor ligands, and an inability to upregulate plasma cell transcription factors. These results build upon previous data demonstrating a role for ATG5 in early B cell development, illustrating its importance in late B cell activation and subsequent plasma cell differentiation.


Assuntos
Diferenciação Celular/imunologia , Proteínas Associadas aos Microtúbulos/metabolismo , Plasmócitos/citologia , Animais , Formação de Anticorpos/imunologia , Antígenos/imunologia , Proteína 5 Relacionada à Autofagia , Epitopos/imunologia , Switching de Imunoglobulina/imunologia , Intestinos/imunologia , Intestinos/parasitologia , Intestinos/patologia , Contagem de Linfócitos , Tecido Linfoide/citologia , Tecido Linfoide/metabolismo , Camundongos , Camundongos Knockout , Proteínas Associadas aos Microtúbulos/deficiência , Nematospiroides dubius/imunologia , Peritônio/citologia , Plasmócitos/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia
19.
Am J Hematol ; 87 Suppl 1: S108-12, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22473489

RESUMO

Testing for hereditary thrombophilia typically includes tests for activated protein C resistance (APC-R) and/or factor V Leiden, protein C, protein S, antithrombin, and prothrombin G20210A. New options for these assays have become available in recent years, with different advantages and disadvantages among the currently available methods. Potential interferences for each assay type are discussed, including lupus anticoagulants, heparin, warfarin, direct thrombin inhibitors (such as argatroban, dabigatran, hirudin, or bivalirudin), rivaroxaban, factor deficiencies or elevations, factor V Leiden, and specific mutations that the assay(s) might not be able to detect. Causes of acquired deficiencies are also described, as these must be carefully excluded before diagnosing a hereditary deficiency of protein C, protein S, or antithrombin.


Assuntos
Anticoagulantes/uso terapêutico , Antitrombinas/uso terapêutico , Proteínas Sanguíneas/genética , Proteínas Sanguíneas/metabolismo , Testes Hematológicos/métodos , Trombofilia , Humanos , Mutação , Trombofilia/sangue , Trombofilia/diagnóstico , Trombofilia/tratamento farmacológico , Trombofilia/genética
20.
PLoS One ; 6(7): e21627, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21765899

RESUMO

Non-productive antigen receptor genes with frame shifts generated during the assembly of these genes are found in many mature lymphocytes. Transcripts from these genes have premature termination codons (PTCs) and could encode truncated proteins if they are not either inactivated or destroyed by nonsense-mediated decay (NMD). In mammalian cells, NMD can be activated by pathways that rely on the presence of an intron downstream of the PTC; however, NMD can also be activated by pathways that do not rely on these downstream introns, and pathways independent of NMD can inactivate PTC-containing transcripts. Here, through the generation and analysis of mice with gene-targeted modifications of the endogenous T cell receptor beta (Tcrb) locus, we demonstrate that in T cells in vivo, optimal clearance of PTC-containing Tcrb transcripts depends on the presence of an intron downstream of the PTC.


Assuntos
Estabilidade de RNA/genética , Fases de Leitura/genética , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Alelos , Animais , Códon sem Sentido/genética , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Linfócitos T/citologia , Linfócitos T/imunologia
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