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1.
Immunity ; 52(2): 257-274.e11, 2020 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-32049053

RESUMO

Genetics is a major determinant of susceptibility to autoimmune disorders. Here, we examined whether genome organization provides resilience or susceptibility to sequence variations, and how this would contribute to the molecular etiology of an autoimmune disease. We generated high-resolution maps of linear and 3D genome organization in thymocytes of NOD mice, a model of type 1 diabetes (T1D), and the diabetes-resistant C57BL/6 mice. Multi-enhancer interactions formed at genomic regions harboring genes with prominent roles in T cell development in both strains. However, diabetes risk-conferring loci coalesced enhancers and promoters in NOD, but not C57BL/6 thymocytes. 3D genome mapping of NODxC57BL/6 F1 thymocytes revealed that genomic misfolding in NOD mice is mediated in cis. Moreover, immune cells infiltrating the pancreas of humans with T1D exhibited increased expression of genes located on misfolded loci in mice. Thus, genetic variation leads to altered 3D chromatin architecture and associated changes in gene expression that may underlie autoimmune pathology.


Assuntos
Cromatina/metabolismo , Diabetes Mellitus Tipo 1/genética , Predisposição Genética para Doença/genética , Timócitos/patologia , Animais , Fator de Ligação a CCCTC/metabolismo , Mapeamento Cromossômico , Diabetes Mellitus Tipo 1/patologia , Epigênese Genética , Expressão Gênica , Loci Gênicos/genética , Variação Genética , Genoma/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Pâncreas/patologia , Sequências Reguladoras de Ácido Nucleico
2.
Sci Rep ; 10(1): 1378, 2020 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-31992777

RESUMO

Previous work showed that the thymus can be infected by RNA viruses as HIV and HTLV-1. We thus hypothesized that the thymus might also be infected by the Zika virus (ZIKV). Herein we provide compelling evidence that ZIKV targets human thymic epithelial cells (TEC) in vivo and in vitro. ZIKV-infection enhances keratinization of TEC, with a decrease in proliferation and increase in cell death. Moreover, ZIKV modulates a high amount of coding RNAs with upregulation of genes related to cell adhesion and migration, as well as non-coding genes including miRNAs, circRNAs and lncRNAs. Moreover, we observed enhanced attachment of lymphoblastic T-cells to infected TEC, as well as virus transfer to those cells. Lastly, alterations in thymuses from babies congenitally infected were seen, with the presence of viral envelope protein in TEC. Taken together, our data reveals that the thymus, particularly the thymic epithelium, is a target for the ZIKV with changes in the expression of molecules that are relevant for interactions with developing thymocytes.


Assuntos
Células Epiteliais , Timócitos , Timo , Tropismo Viral , Infecção por Zika virus , Zika virus/fisiologia , Animais , Chlorocebus aethiops , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Células Epiteliais/virologia , Epitélio/metabolismo , Epitélio/patologia , Epitélio/virologia , Humanos , Timócitos/metabolismo , Timócitos/patologia , Timócitos/virologia , Timo/metabolismo , Timo/patologia , Timo/virologia , Células Vero , Infecção por Zika virus/metabolismo , Infecção por Zika virus/patologia
3.
Scand J Immunol ; 91(3): e12838, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31630413

RESUMO

To evaluate the levels of recent thymic emigrant (RTE) CD4+ T cells in HIV-infected children and to explore the associations among their frequency, antiretroviral treatment (ART) adherence, and CD4+ T cell restoration. The group evaluated comprised 85 HIV-infected patients classified as subjects with moderate or severe immunosuppression or as those with no evidence of immunosuppression. To evaluate the association between the frequency of RTE CD4+ T cells and ART adherence, 23 of the 85 patients were evaluated at two different time points during a one-year follow-up period. Children with severe immunosuppression had lower frequencies of RTE CD4+ T cells compared with children without evidence of immunosuppression (P < .001). The frequency of RTE CD4+ T cells in children with a high rate of adherence was significantly higher (P < .05) than that observed among those with suboptimal adherence. The latter group presented with infectious intercurrences on admission that decreased after initiation of treatment along with improved CD4+ and RTE naïve CD4+ T cells counts. The adequate ART adherence is essential for immune reconstitution, which might be reflected by the levels of RTE CD4+ T cells.


Assuntos
Contagem de Linfócito CD4 , Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Timócitos/imunologia , Adolescente , Terapia Antirretroviral de Alta Atividade , Biomarcadores , Linfócitos T CD4-Positivos/metabolismo , Movimento Celular , Criança , Pré-Escolar , Feminino , Infecções por HIV/tratamento farmacológico , Humanos , Imunofenotipagem , Lactente , Masculino , Adesão à Medicação , Timócitos/metabolismo , Resultado do Tratamento , Carga Viral
4.
Scand J Immunol ; 91(3): e12853, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31793005

RESUMO

What is the evolutionary mechanism for the TCR-MHC-conserved interaction? We extend Dembic's model (Dembic Z. In, Scand J Immunol e12806, 2019) of thymus positive selection for high-avidity anti-self-MHC Tregs among double (CD4 + CD8+)-positive (DP) developing thymocytes. This model is based on competition for self-MHC (+ Pep) complexes presented on cortical epithelium. Such T cells exit as CD4 + CD25+FoxP3 + thymic-derived Tregs (tTregs). The other positively selected DP T cells are then negatively selected on medulla epithelium removing high-avidity anti-self-MHC + Pep as T cells commit to CD4 + or CD8 + lineages. The process is likened to the competitive selection and affinity maturation in Germinal Centre for the somatic hypermutation (SHM) of rearranged immunoglobulin (Ig) variable region (V[D]Js) of centrocytes bearing antigen-specific B cell receptors (BCR). We now argue that the same direct SHM processes for TCRs occur in post-antigenic Germinal Centres, but now occurring in peripheral pTregs. This model provides a potential solution to a long-standing problem previously recognized by Cohn and others (Cohn M, Anderson CC, Dembic Z. In, Scand J Immunol e12790, 2019) of how co-evolution occurs of species-specific MHC alleles with the repertoire of their germline TCR V counterparts. We suggest this is not by 'blind', slow, and random Darwinian natural selection events, but a rapid structured somatic selection vertical transmission process. The pTregs bearing somatic TCR V mutant genes then, on arrival in reproductive tissues, can donate their TCR V sequences via soma-to-germline feedback as discussed in this journal earlier. (Steele EJ, Lindley RA. In, Scand J Immunol e12670, 2018) The high-avidity tTregs also participate in the same process to maintain a biased, high-avidity anti-self-MHC germline V repertoire.


Assuntos
Alelos , Complexo Principal de Histocompatibilidade/genética , Complexo Principal de Histocompatibilidade/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Animais , Diferenciação Celular , Seleção Clonal Mediada por Antígeno , Evolução Molecular , Centro Germinativo/citologia , Centro Germinativo/imunologia , Centro Germinativo/metabolismo , Humanos , Mamíferos , Mutação , Ligação Proteica , Receptores de Antígenos de Linfócitos T/genética , Timócitos/imunologia , Timócitos/metabolismo
5.
Int J Mol Sci ; 21(1)2019 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-31861667

RESUMO

Previous studies have reported the up-regulation of the two-pore domain K+ channel K2P5.1 in the CD4+ T cells of patients with multiple sclerosis (MS) and rheumatoid arthritis (RA), as well as in a mouse model of inflammatory bowel disease (IBD). However, the mechanisms underlying this up-regulation remain unclear. Inflammation-associated hypoxia is involved in the pathogenesis of autoimmune diseases, such as IBD, MS, and RA, and T cells are exposed to a hypoxic environment during their recruitment from inflamed tissues to secondary lymphoid tissues. We herein investigated whether inflammation-associated hypoxia is attributable to the increased expression and activity of K2P5.1 in the splenic CD4+ T cells of chemically-induced IBD model mice. Significant increases in hypoxia-inducible factor (HIF)-1α transcripts and proteins were found in the splenic CD4+ T cells of the IBD model. In the activated splenic CD4+ T cells, hypoxia (1.5% O2) increased K2P5.1 expression and activity, whereas a treatment with the HIF inhibitor FM19G11 but not the selective HIF-2 inhibitor exerted the opposite effect. Hypoxia-exposed K2P5.1 up-regulation was also detected in stimulated thymocytes and the mouse T-cell line. The class III histone deacetylase sirtuin-1 (SIRT1) is a downstream molecule of HIF-1α signaling. We examined the effects of the SIRT1 inhibitor NCO-01 on K2P5.1 transcription in activated CD4+ T cells, and we found no significant effects on the K2P5.1 transcription. No acute compensatory responses of K2P3.1-K2P5.1 up-regulation were found in the CD4+ T cells of the IBD model and the hypoxia-exposed T cells. Collectively, these results suggest a mechanism for K2P5.1 up-regulation via HIF-1 in the CD4+ T cells of the IBD model.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Doenças Inflamatórias Intestinais/genética , Canais de Potássio de Domínios Poros em Tandem/genética , Animais , Benzamidas/farmacologia , Hipóxia Celular , Linhagem Celular , Sulfato de Dextrana/efeitos adversos , Modelos Animais de Doenças , Regulação da Expressão Gênica , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Doenças Inflamatórias Intestinais/induzido quimicamente , Doenças Inflamatórias Intestinais/metabolismo , Camundongos , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Sirtuína 1/genética , Sirtuína 1/metabolismo , Timócitos/citologia , Timócitos/metabolismo
6.
Nat Commun ; 10(1): 5690, 2019 12 19.
Artigo em Inglês | MEDLINE | ID: mdl-31857584

RESUMO

In the thymus, the T lymphocyte repertoire is purged of a substantial portion of highly self-reactive cells. This negative selection process relies on the strength of TCR-signaling in response to self-peptide-MHC complexes, both in the cortex and medulla regions. However, whether cytokine-signaling contributes to negative selection remains unclear. Here, we report that, in the absence of Transforming Growth Factor beta (TGF-ß) signaling in thymocytes, negative selection is significantly impaired. Highly autoreactive thymocytes first escape cortical negative selection and acquire a Th1-like-phenotype. They express high levels of CXCR3, aberrantly accumulate at the cortico-medullary junction and subsequently fail to sustain AIRE expression in the medulla, escaping medullary negative selection. Highly autoreactive thymocytes undergo an atypical maturation program, substantially accumulate in the periphery and induce multiple organ-autoimmune-lesions. Thus, these findings reveal TGF-ß in thymocytes as crucial for negative selection with implications for understanding T cell self-tolerance mechanisms.


Assuntos
Transdução de Sinais , Timócitos/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Autoimunidade , Células da Medula Óssea/metabolismo , Diferenciação Celular , Células Epiteliais/metabolismo , Camundongos Knockout , Modelos Biológicos , Ligante RANK/metabolismo , Timócitos/citologia
7.
Elife ; 82019 12 23.
Artigo em Inglês | MEDLINE | ID: mdl-31868579

RESUMO

Autoreactive thymocytes are eliminated during negative selection in the thymus, a process important for establishing self-tolerance. Thymic phagocytes serve to remove dead thymocytes, but whether they play additional roles during negative selection remains unclear. Here, using a murine thymic slice model in which thymocytes undergo negative selection in situ, we demonstrate that phagocytosis promotes negative selection, and provide evidence for the escape of autoreactive CD8 T cells to the periphery when phagocytosis in the thymus is impaired. We also show that negative selection is more efficient when the phagocyte also presents the negative selecting peptide. Our findings support a model for negative selection in which the death process initiated following strong TCR signaling is facilitated by phagocytosis. Thus, the phagocytic capability of cells that present self-peptides is a key determinant of thymocyte fate.


Assuntos
Morte Celular , Ativação Linfocitária , Fagocitose/fisiologia , Timócitos/metabolismo , Animais , Apresentação do Antígeno , Células da Medula Óssea , Linfócitos T CD8-Positivos/imunologia , Proteínas de Ligação a DNA/genética , Proteínas de Homeodomínio/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Animais , Peptídeos/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Tolerância a Antígenos Próprios , Transdução de Sinais , Timo/imunologia
8.
Adv Immunol ; 144: 1-22, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31699214

RESUMO

During thymocyte development at the double positive stage, thymocytes are subjected to a TCR quality check process termed "thymocyte selection." TCRs with proper binding capabilities to MHC molecules (with self-peptide) are able to transduce cell survival signals and allow the continuing of development to single positive T cells. It has been known that TCRs in DP cells can transduce signals with higher efficiency than peripheral mature T cells, even though they share most of the signaling components. Recent studies have revealed some thymocyte-specific signaling modulators including Themis and Tespa1. The activation of TCR signaling during positive selection results in the activation of several key transcription factors and extensive gene expression change, which has been revealed by newly developed systemic transcriptome analysis tools, and could be used for the evaluation of positive selection process. The fate determination postpositive selection is also governed on the epigenetic level including both DNA methylation and histone modifications.


Assuntos
Epigênese Genética/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Transdução de Sinais/imunologia , Timócitos/imunologia , Fatores de Transcrição/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Animais , Sinalização do Cálcio/imunologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Metilação de DNA/genética , Metilação de DNA/imunologia , Histonas/imunologia , Histonas/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Ativação Linfocitária/imunologia , Camundongos , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais/genética , Timo/citologia , Fatores de Transcrição/genética , Transcriptoma/genética , Transcriptoma/imunologia
9.
Nat Commun ; 10(1): 4698, 2019 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-31619674

RESUMO

T helper 17 (Th17) cells have crucial functions in mucosal immunity and the pathogenesis of several chronic inflammatory diseases. The lineage-specific transcription factor, RORγt, encoded by the RORC gene modulates Th17 polarization and function, as well as thymocyte development. Here we define several regulatory elements at the human RORC locus in thymocytes and peripheral CD4+ T lymphocytes, with CRISPR/Cas9-guided deletion of these genomic segments supporting their role in RORγt expression. Mechanistically, T cell receptor stimulation induces cyclosporine A-sensitive histone modifications and P300/CBP acetylase recruitment at these elements in activated CD4+ T cells. Meanwhile, NFAT proteins bind to these regulatory elements and activate RORγt transcription in cooperation with NF-kB. Our data thus demonstrate that NFAT specifically regulate RORγt expression by binding to the RORC locus and promoting its permissive conformation.


Assuntos
Regulação da Expressão Gênica , Fatores de Transcrição NFATC/metabolismo , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Elementos Reguladores de Transcrição/genética , Células Th17/metabolismo , Timócitos/metabolismo , Ativação Transcricional , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/metabolismo , Sistemas CRISPR-Cas , Linhagem da Célula , Citometria de Fluxo , Células HEK293 , Código das Histonas , Humanos , Células Jurkat , Células Th17/citologia , Timócitos/citologia , Fatores de Transcrição de p300-CBP/metabolismo
10.
Mol Immunol ; 114: 600-611, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31539668

RESUMO

In this work, we demonstrate that adhesion between medullary thymic epithelial cells (mTECs) and thymocytes is controlled by miRNAs. Adhesion between mTECs and developing thymocytes is essential for triggering negative selection (NS) of autoreactive thymocytes that occurs in the thymus. Immune recognition is mediated by the MHC / TCR receptor, whereas adhesion molecules hold cell-cell interaction stability. Indeed, these processes must be finely controlled, if it is not, it may lead to aggressive autoimmunity. Conversely, the precise molecular genetic control of mTEC-thymocyte adhesion is largely unclear. Here, we asked whether miRNAs would be controlling this process through the posttranscriptional regulation of mRNAs that encode adhesion molecules. For this, we used small interfering RNA to knockdown (KD) Dicer mRNA in vitro in a murine mTEC line. A functional assay with fresh murine thymocytes co-cultured with mTECs showed that single-positive (SP) CD4 and CD8 thymocyte adhesion was increased after Dicer KD and most adherent subtype was CD8 SP cells. Analysis of broad mTEC transcriptional expression showed that Dicer KD led to the modulation of 114 miRNAs and 422 mRNAs, including those encoding cell adhesion or extracellular matrix proteins, such as Lgals9, Lgals3pb, Tnc and Cd47. Analysis of miRNA-mRNA networks followed by miRNA mimic transfection showed that these mRNAs are under the control of miR-181b-5p and miR-30b*, which may ultimately control mTEC-thymocyte adhesion. The expression of CD80 surface marker in mTECs was increased after Dicer KD following thymocyte adhesion. This indicates the existence of new mechanisms in mTECs that involve the synergistic action of thymocyte adhesion and regulatory miRNAs.


Assuntos
Adesão Celular/imunologia , Células Epiteliais/imunologia , MicroRNAs/imunologia , Timócitos/imunologia , Timo/imunologia , Animais , Antígeno B7-1/imunologia , Biomarcadores/sangue , Diferenciação Celular/imunologia , Feminino , Regulação da Expressão Gênica/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Antígenos de Linfócitos T/imunologia , Tolerância a Antígenos Próprios/imunologia , Fatores de Transcrição/imunologia
11.
Immunobiology ; 224(5): 614-624, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31427114

RESUMO

Thymus-derived T regulatory (tTregs) cells play a crucial role in the maintenance of tolerance and immune homeostasis. Mechanisms and factors regulating tTreg development and function are widely investigated, but to a large degree still remain unclear. Our previous findings demonstrated that, in physiological conditions, the development and suppressive function of tTregs demonstrated day/night rhythmicity, which correlated with the concentration of plasma corticosterone and the expression of glucocorticoid receptors. In this study we ask whether synthetic glucocorticoids commonly used to inhibit excessive activity of the immune system, can modulate the development and suppressive function of tTregs in vivo depending on the time of administration. Young C57BL/6 male and female mice were injected intraperitoneally with a single dose of dexamethasone at two time points of the day: 7.00-8.00 a.m. and 7.00-8.00 p.m. The experimental can be used to indicate on the potentially expected positive or adverse side effects and can constitute also a good model for the assessment of the effects of long-term therapy. The results of our studies demonstrated the increase of the percentage of tTregs at both time points in male mice, but only in the evening in females. The suppressive activity of tTregs increased independently on the day time of in female mice, but in the morning only in males. We concluded that in the condition of dexamethasone supplementation, the elevated suppressive potential of tTregs is balanced by the induction apoptosis in order to prevent excessive suppression.


Assuntos
Dexametasona/farmacologia , Fotoperíodo , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/fisiologia , Timócitos/efeitos dos fármacos , Timócitos/fisiologia , Animais , Apoptose , Biomarcadores , Diferenciação Celular , Dexametasona/administração & dosagem , Feminino , Glucocorticoides/sangue , Glucocorticoides/metabolismo , Imunofenotipagem , Masculino , Camundongos , Fenótipo , Receptores de Glucocorticoides/metabolismo , Linfócitos T Reguladores/citologia , Timócitos/citologia
12.
Proc Natl Acad Sci U S A ; 116(37): 18537-18543, 2019 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-31451631

RESUMO

Deletion or Treg cell differentiation are alternative fates of autoreactive MHCII-restricted thymocytes. How these different modes of tolerance determine the size and composition of polyclonal cohorts of autoreactive T cells with shared specificity is poorly understood. We addressed how tolerance to a naturally expressed autoantigen of the central nervous system shapes the CD4 T cell repertoire. Specific cells in the tolerant peripheral repertoire either were Foxp3+ or displayed anergy hallmarks and, surprisingly, were at least as frequent as in the nontolerant repertoire. Despite this apparent lack of deletional tolerance, repertoire inventories uncovered that some T cell receptors (TCRs) were lost from the CD4 T cell pool, whereas others mediated Treg cell differentiation. The antigen responsiveness of these TCRs supported an affinity model of central tolerance. Importantly, the contribution of different diverter TCRs to the nascent thymic Treg cell population reflected their antigen reactivity rather than their frequency among precursors. This reveals a multilayered TCR hierarchy in CD4 T cell tolerance that separates deleted and diverted TCRs and assures that the Treg cell compartment is filled with cells of maximal permissive antigen reactivity.


Assuntos
Autoantígenos/imunologia , Diferenciação Celular/imunologia , Deleção Clonal/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Animais , Autoantígenos/genética , Autoantígenos/metabolismo , Linhagem da Célula/genética , Linhagem da Célula/imunologia , Sistema Nervoso Central/imunologia , Sistema Nervoso Central/metabolismo , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/imunologia , Fatores de Transcrição Forkhead/metabolismo , Rearranjo Gênico do Linfócito T/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Ativação Linfocitária , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Proteína Proteolipídica de Mielina/genética , Proteína Proteolipídica de Mielina/imunologia , Proteína Proteolipídica de Mielina/metabolismo , Receptores de Antígenos de Linfócitos T/genética , Linfócitos T Reguladores/metabolismo , Timócitos/fisiologia
13.
Nat Immunol ; 20(10): 1381-1392, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31451788

RESUMO

Proliferation is tightly regulated during T cell development, and is limited to immature CD4-CD8- thymocytes. The major proliferative event is initiated at the 'ß-selection' stage following successful rearrangement of Tcrß, and is triggered by and dependent on concurrent signaling by Notch and the pre-T cell receptor (TCR); however, it is unclear how these signals cooperate to promote cell proliferation. Here, we found that ß-selection-associated proliferation required the combined activity of two Skp-cullin-F-box (SCF) ubiquitin ligase complexes that included as substrate recognition subunits the F-box proteins Fbxl1 or Fbxl12. Both SCF complexes targeted the cyclin-dependent kinase inhibitor Cdkn1b for polyubiquitination and proteasomal degradation. We found that Notch signals induced the transcription of Fbxl1, whereas pre-TCR signals induced the transcription of Fbxl12. Thus, concurrent Notch and pre-TCR signaling induced the expression of two genes, Fbxl1 and Fbxl12, whose products functioned identically but additively to promote degradation of Cdkn1b, cell cycle progression, and proliferation of ß-selected thymocytes.


Assuntos
Proteínas F-Box/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Receptores Notch/metabolismo , Proteínas Ligases SKP Culina F-Box/metabolismo , Linfócitos T/fisiologia , Timócitos/fisiologia , Animais , Diferenciação Celular , Proliferação de Células , Seleção Clonal Mediada por Antígeno , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Proteínas F-Box/genética , Regulação da Expressão Gênica , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T , Camundongos , Camundongos Endogâmicos C57BL , Receptor Cross-Talk , Transdução de Sinais
14.
Immunohorizons ; 3(8): 352-367, 2019 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-31387873

RESUMO

NKAP and HDAC3 are critical for T cell maturation. NKAP and HDAC3 physically associate, and a point mutation in NKAP, NKAP(Y352A), abrogates this interaction. To evaluate the significance of NKAP and HDAC3 association in T cell maturation, transgenic mice were engineered for cre-mediated endogenous NKAP gene deletion coupled to induction of NKAP(Y352A) or a wild type (WT) control transgene, NKAP(WT), in double positive thymocytes or regulatory T cells (Tregs). T cell maturation was normal in mice with endogenous NKAP deletion coupled to NKAP(WT) induction. However, severe defects occurred in T cell and Treg maturation and in iNKT cell development when NKAP(Y352A) was induced, recapitulating NKAP deficiency. Conventional T cells expressing NKAP(Y352A) failed to enter the long-term T cell pool, did not produce cytokines, and remained complement susceptible, whereas Tregs expressing NKAP(Y352A) were eliminated as recent thymic emigrants leading to lethal autoimmunity. Overall, these results demonstrate the significance of NKAP-HDAC3 association in T cells.


Assuntos
Diferenciação Celular/fisiologia , Histona Desacetilases/metabolismo , Células T Matadoras Naturais/metabolismo , Proteínas Repressoras/metabolismo , Linfócitos T Reguladores/metabolismo , Timócitos/metabolismo , Animais , Autoimunidade/genética , Células Cultivadas , Ativação do Complemento , Complemento C3/imunologia , Citocinas/metabolismo , Feminino , Técnicas de Introdução de Genes , Técnicas de Inativação de Genes , Peroxidação de Lipídeos/genética , Masculino , Camundongos , Camundongos Knockout , Mutação Puntual , Proteínas Repressoras/deficiência , Proteínas Repressoras/genética , Timo/citologia
15.
Nat Immunol ; 20(9): 1244-1255, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31431722

RESUMO

Mucosal-associated invariant T cells (MAIT cells) recognize the microbial metabolite 5-(2-oxopropylideneamino)-6-D-ribitylaminouracil (5-OP-RU) presented by the MHC class Ib molecule, MR1. MAIT cells acquire effector functions during thymic development, but the mechanisms involved are unclear. Here we used single-cell RNA-sequencing to characterize the developmental path of 5-OP-RU-specific thymocytes. In addition to the known MAIT1 and MAIT17 effector subsets selected on bone-marrow-derived hematopoietic cells, we identified 5-OP-RU-specific thymocytes that were selected on thymic epithelial cells and differentiated into CD44- naive T cells. MAIT cell positive selection required signaling through the adapter, SAP, that controlled the expression of the transcription factor, ZBTB16. Pseudotemporal ordering of single cells revealed transcriptional trajectories of 5-OP-RU-specific thymocytes selected on either thymic epithelial cells or hematopoietic cells. The resulting model illustrates T cell lineage decisions.


Assuntos
Linhagem da Célula/imunologia , Células T Invariáveis Associadas à Mucosa/citologia , Células T Invariáveis Associadas à Mucosa/imunologia , Ribitol/análogos & derivados , Timócitos/citologia , Timócitos/imunologia , Uracila/análogos & derivados , Animais , Sequência de Bases , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Receptores de Hialuronatos/metabolismo , Ativação Linfocitária/imunologia , Camundongos , Camundongos Knockout , Antígenos de Histocompatibilidade Menor/metabolismo , Proteína com Dedos de Zinco da Leucemia Promielocítica/biossíntese , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Ribitol/imunologia , Análise de Sequência de RNA , Família de Moléculas de Sinalização da Ativação Linfocitária/metabolismo , Timo/citologia , Timo/imunologia , Uracila/imunologia
16.
Food Chem Toxicol ; 133: 110748, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31377140

RESUMO

Hexavalent chromium raises high concern because of its wide industrial applications and reported toxicity. Long-term (135 days) oral exposure of Wistar rats to chromium in the form of K2Cr2O7 (exposed group~20 mg/kg/day) led to a decrease in thymus mass and thymocytes' number and caused structural and functional changes in the lymph nodes and spleen, namely lymphoreticular hyperplasia and plasmocytic macrophage transformation. Programmed cell death was increased in both thymocytes and splenocytes and decreased in lymphocytes in the T-zones of spleen and lymph nodes. Moreover, Cr (VI) administration decreased myeloid cells' and neutrophils' number, while it increased lymphoid and erythroid cells' number in bone marrow. Cr (VI) immune system effects seem to be related to oxidative stress induction, as depicted by the increased levels of diene conjugates and malondialdehyde in the spleen and liver and by the decreased activity of catalase and superoxide dismutase in rats' erythrocytes. In addition, exposure to Cr (VI) decreased copper, nickel and iron concentrations in blood and liver, while Cr levels in blood, spleen and liver were increased, as expected. The observed changes in the series of immunological parameters studied contribute to the development of new approaches for the prevention of low level Cr exposure toxicity.


Assuntos
Cromo/toxicidade , Linfonodos/efeitos dos fármacos , Baço/efeitos dos fármacos , Timo/efeitos dos fármacos , Animais , Peso Corporal/efeitos dos fármacos , Imuno-Histoquímica , Linfonodos/metabolismo , Contagem de Linfócitos , Masculino , Tamanho do Órgão/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Ratos Wistar , Baço/metabolismo , Linfócitos T/metabolismo , Timócitos/metabolismo , Timo/metabolismo , Timo/patologia
17.
Aquat Toxicol ; 214: 105253, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31352076

RESUMO

Diclofop-methyl (DM) is widely used in agriculture and may lead to serious toxicity. However, a limited number of studies have been performed to evaluate the toxicity of DM in the immune and nervous systems of animals. Here, we utilized a good vertebrate model, zebrafish, to evaluate the toxicity of DM during the developmental process. Exposure of zebrafish embryos to 0.1, 0.3 and 0.5 mg/l DM from 6 h post fertilization (hpf) to 72 hpf induced developmental abnormalities, such as shorter body lengths and yolk sac edemas. The number of immune cells in zebrafish larvae was significantly reduced, but the inflammatory response was not influenced by DM treatment. The expression of immune-related genes were downregulated and the levels of oxidative stress were upregulated by DM exposure. Moreover, locomotor behaviors were inhibited by DM exposure. Therefore, our results suggest that DM has the potential to induce immunotoxicity and cause behavioral changes in zebrafish larvae. This study provides new evidence of the influence of DM exposure on aquatic ecosystems.


Assuntos
Comportamento Animal/efeitos dos fármacos , Embrião não Mamífero/imunologia , Exposição Ambiental , Éteres Difenil Halogenados/toxicidade , Peixe-Zebra/embriologia , Animais , Encéfalo/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Larva/efeitos dos fármacos , Larva/metabolismo , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Timócitos/efeitos dos fármacos , Timócitos/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética , Poluentes Químicos da Água/toxicidade , Peixe-Zebra/genética
18.
Mol Biol Rep ; 46(4): 4443-4452, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31302805

RESUMO

Containing high concentration of vanadium served by the volcanic bedrock, Jeju ground water has long been known for various implicit health benefits including immune-promotion. Exposure to stress has been reported to be associated with immunosuppression such as reducing lymphocyte population or antibody production due to stress hormones. In this study, we aimed at evaluating the effects of Jeju ground water on chronically stressed mice. C57BL/6 mice were subjected to various stressors such as restraint stress, water swimming stress, heat stress, acoustic stress, and Jeju ground water was supplied for 28 days with two different concentrations, S1 (vanadium 15-20 µg/l, pH 8.3) and S2 (vanadium 20-25 µg/l, pH 8.5). Treatment with Jeju ground water increased CD4+CD8- or CD4-CD8+ single-positive thymocytes. It also increased the proliferation of splenocytes and the populations of CD4+ T cells, CD45R/B220+ B cells, CD11b+ macrophages or Gr-1+ granulocytes in spleen. In addition, the production of IgG was increased in chronically stressed mice by treatment with Jeju ground water. These results suggest vanadium-rich Jeju ground water may be helpful in T cell development in thymus and immune cell proliferation and its function in spleen against chronic stress.


Assuntos
Linfócitos T/efeitos dos fármacos , Timócitos/efeitos dos fármacos , Vanádio/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Feminino , Água Subterrânea/química , Linfócitos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Baço/imunologia , Estresse Fisiológico/fisiologia , Timo/imunologia , Vanádio/fisiologia
19.
Proc Natl Acad Sci U S A ; 116(25): 12422-12427, 2019 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-31152132

RESUMO

The development of thymocytes to mature T cells in the thymus is tightly controlled by cellular selection, in which only a small fraction of thymocytes equipped with proper quality of TCRs progress to maturation. It is pivotal to protect the survival of the few T cells, which pass the selection. However, the signaling events, which safeguard the cell survival in thymus, are not totally understood. In this study, protein Ser/Thr phosphorylation in thymocytes undergoing positive selection is profiled by mass spectrometry. The results revealed large numbers of dephosphorylation changes upon T cell receptor (TCR) activation during positive selection. Subsequent substrate analysis pinpointed protein phosphatase 2A (PP2A) as the enzyme responsible for the dephosphorylation changes in developing thymocytes. PP2A catalytic subunit α (Ppp2ca) deletion in the T cell lineage in Ppp2ca flox/flox-Lck-Cre mice (PP2A cKO) displayed dysregulated dephosphorylation of apoptosis-related proteins in double-positive (DP) cells and caused substantially decreased numbers of DP CD4+ CD8+ cells. Increased levels of apoptosis in PP2A cKO DP cells were found to underlie aberrant thymocyte development. Finally, the defective thymocyte development in PP2A cKO mice could be rescued by either Bcl2 transgene expression or by p53 knockout. In summary, our work reveals an essential role of PP2A in promoting thymocyte development through the regulation of cell survival.


Assuntos
Sobrevivência Celular , Proteína Fosfatase 2/metabolismo , Timócitos/citologia , Animais , Apoptose , Proliferação de Células , Genes p53 , Camundongos , Camundongos Knockout , Fosforilação , Proteína Fosfatase 2/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Timócitos/enzimologia
20.
J Immunol ; 203(2): 408-417, 2019 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-31175160

RESUMO

Recent thymic emigrants that fail postpositive selection maturation are targeted by complement proteins. T cells likely acquire complement resistance during maturation in the thymus, a complement-privileged organ. To test this, thymocytes and fresh serum were separately obtained and incubated together in vitro to assess complement deposition. Complement binding decreased with development and maturation. Complement binding decreased from the double-positive thymocyte to the single-positive stage, and within single-positive thymocytes, complement binding gradually decreased with increasing intrathymic maturation. Binding of the central complement protein C3 to wild-type immature thymocytes required the lectin but not the classical pathway. Specifically, MBL2 but not MBL1 was required, demonstrating a unique function for MBL2. Previous studies demonstrated that the loss of NKAP, a transcriptional regulator of T cell maturation, caused peripheral T cell lymphopenia and enhanced complement susceptibility. To determine whether complement causes NKAP-deficient T cell disappearance, both the lectin and classical pathways were genetically ablated. This blocked C3 deposition on NKAP-deficient T cells but failed to restore normal cellularity, indicating that complement contributes to clearance but is not the primary cause of peripheral T cell lymphopenia. Rather, the accumulation of lipid peroxides in NKAP-deficient T cells was observed. Lipid peroxidation is a salient feature of ferroptosis, an iron-dependent nonapoptotic cell death. Thus, wild-type thymocytes naturally acquire the ability to protect themselves from complement targeting by MBL2 with maturation. However, NKAP-deficient immature peripheral T cells remain scarce in complement-deficient mice likely due to ferroptosis.


Assuntos
Diferenciação Celular/imunologia , Complemento C3/imunologia , Lectina de Ligação a Manose/imunologia , Proteínas Repressoras/imunologia , Linfócitos T/imunologia , Animais , Linfopenia/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Timócitos/imunologia , Timo/imunologia , Transcrição Genética/imunologia
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