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1.
J Exp Med ; 2019 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-31537641

RESUMO

Infection of T cells by Epstein-Barr virus (EBV) causes chronic active EBV infection (CAEBV) characterized by T cell lymphoproliferative disorders (T-LPD) of unclear etiology. Here, we identified two homozygous biallelic loss-of-function mutations in PIK3CD and TNFRSF9 in a patient who developed a fatal CAEBV. The mutation in TNFRSF9 gene coding CD137/4-1BB, a costimulatory molecule expressed by antigen-specific activated T cells, resulted in a complete loss of CD137 expression and impaired T cell expansion toward CD137 ligand-expressing cells. Isolated as observed in one sibling, CD137 deficiency resulted in persistent EBV-infected T cells but without clinical manifestations. The mutation in PIK3CD gene that encodes the catalytic subunit p110δ of the PI3K significantly reduced its kinase activity. Deficient T cells for PIK3CD exhibited reduced AKT signaling, while calcium flux, RAS-MAPK activation, and proliferation were increased, suggestive of an imbalance between the PLCγ1 and PI3K pathways. These skewed signals in T cells may sustain accumulation of EBV-infected T cells, a process controlled by the CD137-CD137L pathway, highlighting its critical role in immunity to EBV.

2.
Eur J Immunol ; 49(6): 894-910, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30912587

RESUMO

It is established that iNKT cells are a cell type that require strong TCR signal for their proper development and represent a model for thymic agonist selection. The nature of the signal perceived by iNKT cells promoting their specification is not well understood. To address this question, we analyzed iNKT cell development in relevant TCR Vα14-Jα18 alpha chain transgenic mice (Vα14Tg). In CD4-Vα14Tg mice, where the transgene is driven by CD4 promoter, we identified a block in iNKT cell development at early developmental stages due to a reduced expression of key transcription factors accompanied with a reduced TCR expression levels. This indicates that TCR signal strength control iNKT cell differentiation. Importantly, we found in WT mice that early precursors of iNKT cells express higher TCR levels compared to positively selected precursors of mainstream T cells showing that TCR levels could contribute to the strength of iNKT cell TCR signaling. Overall, our study highlights TCR signal strength associated with a higher TCR density as an important regulator of iNKT cell lineage specification.

4.
J Clin Invest ; 128(7): 3071-3087, 2018 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-29889099

RESUMO

Ikaros/IKZF1 is an essential transcription factor expressed throughout hematopoiesis. IKZF1 is implicated in lymphocyte and myeloid differentiation and negative regulation of cell proliferation. In humans, somatic mutations in IKZF1 have been linked to the development of B cell acute lymphoblastic leukemia (ALL) in children and adults. Recently, heterozygous germline IKZF1 mutations have been identified in patients with a B cell immune deficiency mimicking common variable immunodeficiency. These mutations demonstrated incomplete penetrance and led to haploinsufficiency. Herein, we report 7 unrelated patients with a novel early-onset combined immunodeficiency associated with de novo germline IKZF1 heterozygous mutations affecting amino acid N159 located in the DNA-binding domain of IKZF1. Different bacterial and viral infections were diagnosed, but Pneumocystis jirovecii pneumonia was reported in all patients. One patient developed a T cell ALL. This immunodeficiency was characterized by innate and adaptive immune defects, including low numbers of B cells, neutrophils, eosinophils, and myeloid dendritic cells, as well as T cell and monocyte dysfunctions. Notably, most T cells exhibited a naive phenotype and were unable to evolve into effector memory cells. Functional studies indicated these mutations act as dominant negative. This defect expands the clinical spectrum of human IKZF1-associated diseases from somatic to germline, from haploinsufficient to dominant negative.

5.
J Crohns Colitis ; 2018 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-29788237

RESUMO

Background and Aims: An expanding number of monogenic defects have been identified as causative of severe forms of very early-onset inflammatory bowel diseases (VEO-IBD). The present study aimed at defining how next-generation sequencing (NGS) methods can be used to improve identification of known molecular diagnosis and adapt treatment. Methods: 207 children were recruited in 45 Paediatric centres through an international collaborative network (ESPGHAN GENIUS working group) with a clinical presentation of severe VEO-IBD (n=185) or an anamnesis suggestive of a monogenic disorder (n=22). Patients were divided at inclusion into three phenotypic subsets: predominantly small bowel inflammation, colitis with perianal lesions, and colitis only. Methods to obtain molecular diagnosis included functional tests followed by specific Sanger sequencing, custom-made targeted NGS, and in selected cases whole exome sequencing (WES) of parents-child trios. Genetic findings were validated clinically and/or functionally. Results: Molecular diagnosis was achieved in 66/207 children (32%): 61% with small bowel inflammation, 39% with colitis and perianal lesions and 18% with colitis only. Targeted NGS pinpointed gene mutations causative of atypical presentations and identified large exonic copy number variations previously missed by WES. Conclusions: Our results lead us to propose an optimised diagnostic strategy to identify known monogenic causes of severe IBD.

6.
EMBO Mol Med ; 10(2): 188-199, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29282224

RESUMO

Inherited CTPS1, CD27, and CD70 deficiencies in humans have revealed key factors of T-lymphocyte expansion, a critical prerequisite for an efficient immunity to Epstein-Barr virus (EBV) infection. RASGRP1 is a T-lymphocyte-specific nucleotide exchange factor known to activate the pathway of MAP kinases (MAPK). A deleterious homozygous mutation in RASGRP1 leading to the loss RASGRP1 expression was identified in two siblings who both developed a persistent EBV infection leading to Hodgkin lymphoma. RASGRP1-deficient T cells exhibited defective MAPK activation and impaired proliferation that was restored by expression of wild-type RASGRP1. Similar defects were observed in T cells from healthy individuals when RASGRP1 was downregulated. RASGRP1-deficient T cells also exhibited decreased CD27-dependent proliferation toward CD70-expressing EBV-transformed B cells, a crucial pathway required for expansion of antigen-specific T cells during anti-EBV immunity. Furthermore, RASGRP1-deficient T cells failed to upregulate CTPS1, an important enzyme involved in DNA synthesis. These results show that RASGRP1 deficiency leads to susceptibility to EBV infection and demonstrate the key role of RASGRP1 at the crossroad of pathways required for the expansion of activated T lymphocytes.

7.
J Exp Med ; 214(1): 73-89, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-28011863

RESUMO

Epstein-Barr virus (EBV) infection in humans is a major trigger of malignant and nonmalignant B cell proliferations. CD27 is a co-stimulatory molecule of T cells, and inherited CD27 deficiency is characterized by high susceptibility to EBV infection, though the underlying pathological mechanisms have not yet been identified. In this study, we report a patient suffering from recurrent EBV-induced B cell proliferations including Hodgkin's lymphoma because of a deficiency in CD70, the ligand of CD27. We show that EBV-specific T lymphocytes did not expand properly when stimulated with CD70-deficient EBV-infected B cells, whereas expression of CD70 in B cells restored expansion, indicating that CD70 on B cells but not on T cells is required for efficient proliferation of T cells. CD70 was found to be up-regulated on B cells when activated and during EBV infection. The proliferation of T cells triggered by CD70-expressing B cells was dependent on CD27 and CD3 on T cells. Importantly, CD27-deficient T cells failed to proliferate when stimulated with CD70-expressing B cells. Thus, the CD70-CD27 pathway appears to be a crucial component of EBV-specific T cell immunity and more generally for the immune surveillance of B cells and may be a target for immunotherapy of B cell malignancies.


Assuntos
Ligante CD27/fisiologia , Infecções por Vírus Epstein-Barr/imunologia , Transdução de Sinais/fisiologia , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/fisiologia , Linfócitos B/imunologia , Ligante CD27/deficiência , Ligante CD27/genética , Criança , Códon sem Sentido , Humanos , Ativação Linfocitária , Masculino , Receptores de Antígenos de Linfócitos T/fisiologia , Linfócitos T/imunologia
8.
Eur J Immunol ; 46(9): 2162-74, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-27338553

RESUMO

Invariant NKT (iNKT) cells differentiate in the thymus into three distinct lineages defined by their cytokine and transcription factor expression. Signaling lymphocyte activation molecule (SLAM)-associated protein (SAP) is essential for early stages of iNKT cell development, but its role during terminal differentiation of iNKT1, iNKT2, or iNKT17 cells remains unclear. Taking advantage of SAP-deficient mice expressing a Vα14-Jα18 TCRα transgene, we found that SAP is critical not only for IL-4 production but also for the terminal differentiation of IL-4-producing iNKT2 cells. Furthermore, without SAP, the IL-17 producing subset is expanded, while IFN-γ-producing iNKT1 differentiation is only moderately compromised. Lack of SAP reduced the expression of the transcription factors GATA-3 and promyelocytic leukemia zinc finger, but enhanced the levels of retinoic acid receptor-related orphan receptor γt. In the absence of SAP, lineage commitment was actually shifted toward the emergence of iNKT17 over iNKT2 cells. Collectively, our data unveil a new critical regulatory function for SAP in thymic iNKT cell fate decisions.


Assuntos
Diferenciação Celular/imunologia , Células T Matadoras Naturais/citologia , Células T Matadoras Naturais/metabolismo , Proteína Associada à Molécula de Sinalização da Ativação Linfocitária/metabolismo , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/metabolismo , Animais , Biomarcadores , Células Cultivadas , Imunofenotipagem , Interleucina-17/biossíntese , Interleucina-4/biossíntese , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Fenótipo , Proteína Associada à Molécula de Sinalização da Ativação Linfocitária/deficiência , Proteína Associada à Molécula de Sinalização da Ativação Linfocitária/genética
9.
Nat Immunol ; 17(4): 387-96, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26878112

RESUMO

Activation of natural killer (NK) cells by hematopoietic target cells is controlled by the SLAM family of receptors and by the associated SAP family of adaptors. Here we found that SLAM receptors also enhanced NK cell activation by nonhematopoietic target cells, which lack ligands for SLAM receptors. This function was mediated by SLAMF6, a homotypic SLAM receptor found on NK cells and other hematopoietic cells, and was regulated by SAP adaptors, which uncoupled SLAM receptors from phosphatase SHP-1 and diminished the effect of SLAMF6 on NK cell responsiveness toward nonhematopoietic cells. Thus, in addition to their role in NK cell activation by hematopoietic cells, the SLAM-SAP pathways influence responsiveness toward nonhematopoietic targets by a process akin to NK cell 'education'.


Assuntos
Antígenos CD/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Células Matadoras Naturais/imunologia , Proteína Tirosina Fosfatase não Receptora Tipo 6/imunologia , Receptores de Superfície Celular/imunologia , Transferência Adotiva , Animais , Linhagem Celular Tumoral , Células HeLa , Humanos , Imunidade Inata , Ativação Linfocitária , Melanoma Experimental , Camundongos , Transdução de Sinais , Proteína Associada à Molécula de Sinalização da Ativação Linfocitária , Família de Moléculas de Sinalização da Ativação Linfocitária , Membro 1 da Família de Moléculas de Sinalização da Ativação Linfocitária
11.
Clin Immunol ; 161(2): 103-9, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26187144

RESUMO

Autosomal recessive human ZAP70 deficiency is a rare cause of combined immunodeficiency (CID) characterized by defective CD4 T cells and profound CD8 T cell lymphopenia. Herein, we report two novel patients that extend the molecular genetics, the clinical and functional phenotypes associated with the ZAP70 deficiency. The patients presented as infant-onset CID with severe infections caused by varicella zoster virus and live vaccines. Retrospective TCR excision circle newborn screening was normal in both patients. One patient carried a novel non-sense mutation (p.A495fsX75); the other a previously described misense mutation (p.A507V). In contrast to CD4 T cells, the majority of the few CD8 T cells showed expression of the ZAP70-related tyrosine kinase SYK that correlated with residual TCR signaling including calcium flux and degranulation. Our findings highlight the differential requirements of ZAP70 and SYK during thymic development, peripheral homeostasis as well as effector functions of CD4 and CD8 T cells.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Proteínas Tirosina Quinases/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Imunodeficiência Combinada Severa/imunologia , Proteína-Tirosina Quinase ZAP-70/deficiência , Linfócitos T CD4-Positivos/imunologia , Pré-Escolar , Humanos , Síndromes de Imunodeficiência/imunologia , Lactente , Masculino , Mutação/imunologia , Estudos Retrospectivos , Transdução de Sinais/imunologia , Quinase Syk , Proteína-Tirosina Quinase ZAP-70/imunologia
13.
J Allergy Clin Immunol ; 134(5): 1131-41.e9, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24942515

RESUMO

BACKGROUND: Crohn disease is an inflammatory bowel disease (IBD) with a complex mode of inheritance. Although nucleotide binding and oligomerization domain containing 2 (NOD2) is the strongest risk factor, the cause of Crohn disease remains unknown in the majority of the cases. X-linked inhibitor of apoptosis (XIAP) deficiency causes X-linked lymphoproliferative syndrome type 2. IBD has been reported in some XIAP-deficient patients. OBJECTIVE: We characterize the IBD affecting a large cohort of patients with mutations in XIAP and examine the possible pathophysiologic mechanisms. METHODS: We performed a phenotypical and histologic analysis of the IBD affecting 17 patients with hemizygous mutations in XIAP, including 3 patients identified by screening 83 patients with pediatric-onset IBD. The X chromosome inactivation was analyzed in female carriers of heterozygous XIAP mutations, including 2 adults with IBD. The functional consequences of XIAP deficiency were analyzed. RESULTS: Clinical presentation and histology of IBD in patients with XIAP deficiency overlapped with those of patients with Crohn disease. The age at onset was variable (from 3 months to 41 years), and IBD was severe and difficult to treat. In 2 patients hematopoietic stem cell transplantation fully restored intestinal homeostasis. Monocytes of patients had impaired NOD2-mediated IL-8 and monocyte chemoattractant protein 1 (MCP-1) production, as well as IL-10, in response to NOD2 and Toll-like receptor 2/4 costimulation. Nucleotide binding and oligomerization domain containing 1 (NOD1)-mediated IL-6 and IL-8 production was defective in fibroblasts from XIAP-deficient patients. The 2 heterozygous female carriers of XIAP mutations with IBD displayed abnormal expression of the XIAP mutated allele, resulting in impaired activation of the NOD2 pathway. CONCLUSION: IBD in patients with XIAP deficiency is similar to Crohn disease and is associated with defective NOD2 function in monocytes. Importantly, we report that it is not restricted to male patients because we identified 2 symptomatic female heterozygous carriers of XIAP mutations.


Assuntos
Cromossomos Humanos X , Doença de Crohn , Doenças Genéticas Ligadas ao Cromossomo X , Hemizigoto , Heterozigoto , Transtornos Linfoproliferativos , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/genética , Adolescente , Adulto , Idade de Início , Criança , Pré-Escolar , Cromossomos Humanos X/genética , Cromossomos Humanos X/metabolismo , Estudos de Coortes , Doença de Crohn/sangue , Doença de Crohn/genética , Doença de Crohn/patologia , Citocinas/sangue , Citocinas/genética , Feminino , Doenças Genéticas Ligadas ao Cromossomo X/sangue , Doenças Genéticas Ligadas ao Cromossomo X/genética , Doenças Genéticas Ligadas ao Cromossomo X/patologia , Humanos , Lactente , Transtornos Linfoproliferativos/sangue , Transtornos Linfoproliferativos/genética , Transtornos Linfoproliferativos/patologia , Masculino , Proteína Adaptadora de Sinalização NOD2/genética , Proteína Adaptadora de Sinalização NOD2/metabolismo
14.
Nature ; 510(7504): 288-92, 2014 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-24870241

RESUMO

Lymphocyte functions triggered by antigen recognition and co-stimulation signals are associated with a rapid and intense cell division, and hence with metabolism adaptation. The nucleotide cytidine 5' triphosphate (CTP) is a precursor required for the metabolism of DNA, RNA and phospholipids. CTP originates from two sources: a salvage pathway and a de novo synthesis pathway that depends on two enzymes, the CTP synthases (or synthetases) 1 and 2 (CTPS1 with CTPS2); the respective roles of these two enzymes are not known. CTP synthase activity is a potentially important step for DNA synthesis in lymphocytes. Here we report the identification of a loss-of-function homozygous mutation (rs145092287) in CTPS1 in humans that causes a novel and life-threatening immunodeficiency, characterized by an impaired capacity of activated T and B cells to proliferate in response to antigen receptor-mediated activation. In contrast, proximal and distal T-cell receptor (TCR) signalling events and responses were only weakly affected by the absence of CTPS1. Activated CTPS1-deficient cells had decreased levels of CTP. Normal T-cell proliferation was restored in CTPS1-deficient cells by expressing wild-type CTPS1 or by addition of exogenous CTP or its nucleoside precursor, cytidine. CTPS1 expression was found to be low in resting T cells, but rapidly upregulated following TCR activation. These results highlight a key and specific role of CTPS1 in the immune system by its capacity to sustain the proliferation of activated lymphocytes during the immune response. CTPS1 may therefore represent a therapeutic target of immunosuppressive drugs that could specifically dampen lymphocyte activation.


Assuntos
Carbono-Nitrogênio Ligases/deficiência , Carbono-Nitrogênio Ligases/metabolismo , Ativação Linfocitária , Linfócitos/citologia , Linfócitos B/citologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Complexo CD3/imunologia , Carbono-Nitrogênio Ligases/genética , Proliferação de Células , Pré-Escolar , Citidina Trifosfato/metabolismo , Feminino , Humanos , Síndromes de Imunodeficiência/enzimologia , Síndromes de Imunodeficiência/genética , Lactente , Recém-Nascido , Ativação Linfocitária/genética , Linfócitos/imunologia , Linfócitos/metabolismo , Masculino , Mutação/genética , Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia , Linfócitos T/metabolismo
15.
Blood ; 121(4): 614-23, 2013 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-23223428

RESUMO

Invariant natural killer (iNKT) T cells and mucosal-associated invariant T (MAIT) cells represent peculiar T-lymphocyte subpopulations with innate-like properties that differ from conventional T cells. iNKT are reduced in the primary immunodeficiency caused by mutations in the X-linked inhibitor of apoptosis (XIAP). By studying the mechanism of this depletion, we herein report that iNKT cells exhibit a high susceptibility to apoptosis that is not observed with conventional T cells. Elevated expression of caspases 3 and 7 accounts for the proapoptotic phenotype of iNKT cells, which is inhibited by XIAP although it exerts a moderate effect in conventional T cells. Similarly, MAIT cells exhibit a proapoptotic propensity with elevated expression of activated caspases and are decreased in XIAP-deficient individuals. Knockdown of the transcription factor PLZF/ZBTB-16, which is involved in the effector program of iNKT cells, diminishes their proapoptotic phenotype. Conversely, overexpression of PLZF/ZBTB-16 in conventional T cells leads to a proapoptotic phenotype. Our findings identify a previously unknown pathway of regulation of innate-like T-cell homeostasis depending on XIAP and PLZF. The proapoptotic feature of iNKT cells also gives a reliable explanation of their exhaustion observed in different human conditions including the XIAP immunodeficiency.


Assuntos
Fatores de Transcrição Kruppel-Like/genética , Células T Matadoras Naturais/metabolismo , Subpopulações de Linfócitos T/metabolismo , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/genética , Apoptose/genética , Apoptose/imunologia , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/imunologia , Regulação da Expressão Gênica , Humanos , Fatores de Transcrição Kruppel-Like/imunologia , Contagem de Linfócitos , Células T Matadoras Naturais/imunologia , Fenótipo , Proteína com Dedos de Zinco da Leucemia Promielocítica , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Subpopulações de Linfócitos T/imunologia , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/imunologia
16.
Blood ; 118(2): 252-61, 2011 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-21543760

RESUMO

The present study focuses on a large family with an X-linked immunodeficiency in which there are variable clinical and laboratory phenotypes, including recurrent viral and bacterial infections, hypogammaglobulinemia, Epstein-Barr virus-driven lymphoproliferation, splenomegaly, colitis, and liver disease. Molecular and genetic analyses revealed that affected males were carriers of a hypomorphic hemizygous mutation in XIAP (XIAP(G466X)) that cosegregated with a rare polymorphism in CD40LG (CD40 ligand(G219R)). These genes are involved in the X-linked lymphoproliferative syndrome 2 and the X-linked hyper-IgM syndrome, respectively. Single expression of XIAP(G466X) or CD40L(G219R) had no or minimal effect in vivo, although in vitro, they lead to altered functional activities of their gene products, which suggests that the combination of XIAP and CD40LG mutations contributed to the expression of clinical manifestations observed in affected individuals. Our report of a primary X-linked immunodeficiency of oligogenic origin emphasizes that primary immunodeficiencies are not caused by a single defective gene, which leads to restricted manifestations, but are likely to be the result of an interplay between several genetic determinants, which leads to more variable clinical phenotypes.


Assuntos
Ligante de CD40/genética , Imunodeficiência de Variável Comum/genética , Transtornos Linfoproliferativos/genética , Polimorfismo de Nucleotídeo Único , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/genética , Adolescente , Adulto , Arginina/genética , Criança , Epistasia Genética/fisiologia , Família , Feminino , Genes Ligados ao Cromossomo X , Glutamina/genética , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Linhagem , Polimorfismo de Nucleotídeo Único/fisiologia , Adulto Jovem
17.
Blood ; 117(5): 1522-9, 2011 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-21119115

RESUMO

X-linked lymphoproliferative syndromes (XLP) are primary immunodeficiencies characterized by a particular vulnerability toward Epstein-Barr virus infection, frequently resulting in hemophagocytic lymphohistiocytosis (HLH). XLP type 1 (XLP-1) is caused by mutations in the gene SH2D1A (also named SAP), whereas mutations in the gene XIAP underlie XLP type 2 (XLP-2). Here, a comparison of the clinical phenotypes associated with XLP-1 and XLP-2 was performed in cohorts of 33 and 30 patients, respectively. HLH (XLP-1, 55%; XLP-2, 76%) and hypogammaglobulinemia (XLP-1, 67%; XLP-2, 33%) occurred in both groups. Epstein-Barr virus infection in XLP-1 and XLP-2 was the common trigger of HLH (XLP-1, 92%; XLP-2, 83%). Survival rates and mean ages at the first HLH episode did not differ for both groups, but HLH was more severe with lethal outcome in XLP-1 (XLP-1, 61%; XLP-2, 23%). Although only XLP-1 patients developed lymphomas (30%), XLP-2 patients (17%) had chronic hemorrhagic colitis as documented by histopathology. Recurrent splenomegaly often associated with cytopenia and fever was preferentially observed in XLP-2 (XLP-1, 7%; XLP-2, 87%) and probably represents minimal forms of HLH as documented by histopathology. This first phenotypic comparison of XLP subtypes should help to improve the diagnosis and the care of patients with XLP conditions.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/genética , Transtornos Linfoproliferativos/diagnóstico , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/genética , Adolescente , Adulto , Idoso , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Humanos , Técnicas Imunoenzimáticas , Lactente , Transtornos Linfoproliferativos/genética , Transtornos Linfoproliferativos/terapia , Masculino , Pessoa de Meia-Idade , Mutação/genética , Fenótipo , Estudos Retrospectivos , Proteína Associada à Molécula de Sinalização da Ativação Linfocitária , Taxa de Sobrevida , Adulto Jovem
18.
Life Sci ; 82(9-10): 519-28, 2008 Feb 27.
Artigo em Inglês | MEDLINE | ID: mdl-18215718

RESUMO

Intestinal epithelial cells (IEC) have adapted to the presence of commensal bacteria through a state of tolerance that involves a limited response to lipopolysaccharide (LPS). Low or absent expression of two LPS receptor molecules, the myeloid differentiation (MD)-2 receptor, and toll-like receptor (TLR)4 was suggested to underlie LPS tolerance in IEC. In the present study we performed transfections of TLR4 and MD-2 alone or combined in different IEC lines derived from intestinal cancer (Caco-2, HT-29, and SW837). We found that LPS responsiveness increased more than 100-fold when IEC were transfected with MD-2 alone, but not TLR4. The release of interleukin (IL)-8, but also the expression of cyclooxygenase (Cox-)2 and the related secretion of prostaglandin (PG)E2 were coordinately stimulated by LPS in IEC transfected with MD-2 alone. Supernatants collected from MD-2-transfected IEC supported LPS activation of naïve HT-29, providing additional support to the concept that MD-2 alone endows IEC with LPS responsiveness. LPS responsiveness detected at concentrations as low as 110 pg/ml, and maximal values obtained by 10 ng/ml were clearly beyond those evoked by classical stimuli as IL-1beta. In polarized cells, apical LPS stimulation was markedly more efficient than basolateral. Our data contradict previous opinion that both TLR4 and MD-2 limit IEC response to LPS, and emphasize the prominent role of MD-2 in intestinal immune responses to Gram-negative bacteria.


Assuntos
Células Epiteliais/efeitos dos fármacos , Lipopolissacarídeos/farmacologia , Antígeno 96 de Linfócito/fisiologia , Antracenos/farmacologia , Células CACO-2 , Linhagem Celular Tumoral , Células Cultivadas , Ciclo-Oxigenase 2/biossíntese , Dinoprostona/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Citometria de Fluxo , Expressão Gênica , Células HT29 , Humanos , Imidazóis/farmacologia , Interleucina-8/metabolismo , Intestinos/patologia , Antígeno 96 de Linfócito/genética , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Monócitos/citologia , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , NF-kappa B/metabolismo , Piridinas/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/fisiologia , Transfecção
19.
J Virol ; 81(3): 1297-304, 2007 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17079279

RESUMO

Previous studies demonstrated that the induction of the heat shock protein Hsp70 in response to viral infection is highly specific and differs from one cell to another and for a given virus type. However, no clear consensus exists so far to explain the likely reasons for Hsp70 induction within host cells during viral infection. We show here that upon rotavirus infection of intestinal cells, Hsp70 is indeed rapidly, specifically, and transiently induced. Using small interfering RNA-Hsp70-transfected Caco-2 cells, we observed that Hsp70 silencing was associated with an increased virus protein level and enhanced progeny virus production. Upon Hsp70 silencing, we observed that the ubiquitination of the main rotavirus structural proteins was strongly reduced. In addition, the use of proteasome inhibitors in infected Caco-2 cells was shown to induce an accumulation of structural viral proteins. Together, these results are consistent with a role of Hsp70 in the control of the bioavailability of viral proteins within cells for virus morphogenesis.


Assuntos
Regulação Viral da Expressão Gênica/fisiologia , Proteínas de Choque Térmico HSP70/fisiologia , Rotavirus/fisiologia , Proteínas Virais/metabolismo , Células CACO-2/metabolismo , Células CACO-2/virologia , Proteínas de Choque Térmico HSP70/genética , Humanos , Rotavirus/crescimento & desenvolvimento , Proteínas Virais/biossíntese
20.
J Biol Chem ; 278(39): 37799-809, 2003 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-12855686

RESUMO

In previous work we reported that long term treatment of polarized HT-29 cells by 1-benzyl-2-acetamido-2-deoxy-alpha-d-galactopyranoside (GalNAcalpha-O-bn) induced undersialylation and intracellular distribution of apical glycoproteins such as dipeptidyl peptidase IV (DPP-IV), and we suggested therefore that sialylation could act as an apical targeting signal. In this work, the apical direct biosynthetic route was studied after transfection of polarized enterocyte-like HT-29 5M12 cloned cells with a murine cDNA coding for a soluble form of DPP-IV, which was secreted into the apical medium. A 24-h treatment of transfected cells by GalNAcalpha-O-bn markedly inhibited the apical secretion and the sialylation of this soluble murine DPP-IV, which became blocked inside the cell. A similar short GalNAcalpha-O-bn treatment also induced an intracellular distribution of both endogenous transmembrane DPP-IV and proteins involved in the regulation of the apical trafficking such as the apical t-SNARE syntaxin-3 and the raft-associated protein annexin XIIIb, whereas the basolateral t-SNARE syntaxin-4 kept its normal localization. These apical membrane proteins moved efficiently from trans-Golgi network to apical carrier vesicles but failed to be transported from carrier vesicles to the apical plasma membrane. Isolation of membrane microdomains showed that GalNAcalpha-O-bn induced the formation of abnormal lipid-rich microdomains in comparison to normal rafts, as shown by their lower buoyant density and their depletion in annexin XIIIb. In conclusion, GalNAcalpha-O-bn blocks the anterograde traffic to the apical surface of polarized HT-29 cells at the transport level or docking/fusion level of carrier vesicles.


Assuntos
Galactose/análogos & derivados , Galactose/farmacologia , Proteínas de Transporte Vesicular , Animais , Anexinas/metabolismo , Polaridade Celular , Dipeptidil Peptidase 4/metabolismo , Glicosilação , Células HT29 , Humanos , Microdomínios da Membrana/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Transporte Proteico , Proteínas Qa-SNARE , Proteínas SNARE
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