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1.
Clin Proteomics ; 19(1): 23, 2022 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-35790914

RESUMO

BACKGROUND: Multiple sclerosis (MS) is an autoimmune, neurodegenerative disorder with a strong genetic component that acts in a complex interaction with environmental factors for disease development. CD4+ T cells are pivotal players in MS pathogenesis, where peripherally activated T cells migrate to the central nervous system leading to demyelination and axonal degeneration. Through a proteomic approach, we aim at identifying dysregulated pathways in activated T cells from MS patients as compared to healthy controls. METHODS: CD4+ T cells were purified from peripheral blood from MS patients and healthy controls by magnetic separation. Cells were left unstimulated or stimulated in vitro through the TCR and costimulatory CD28 receptor for 24 h prior to sampling. Electrospray liquid chromatography-tandem mass spectrometry was used to measure protein abundances. RESULTS: Upon T cell activation the abundance of 1801 proteins was changed. Among these proteins, we observed an enrichment of proteins expressed by MS-susceptibility genes. When comparing protein abundances in T cell samples from healthy controls and MS patients, 18 and 33 proteins were differentially expressed in unstimulated and stimulated CD4+ T cells, respectively. Moreover, 353 and 304 proteins were identified as proteins exclusively induced upon T cell activation in healthy controls and MS patients, respectively and dysregulation of the Nur77 pathway was observed only in samples from MS patients. CONCLUSIONS: Our study highlights the importance of CD4+ T cell activation for MS, as proteins that change in abundance upon T cell activation are enriched for proteins encoded by MS susceptibility genes. The results provide evidence for proteomic disturbances in T cell activation in MS, and pinpoint to dysregulation of the Nur77 pathway, a biological pathway known to limit aberrant effector T cell responses.

2.
J Biol Chem ; 287(27): 22927-37, 2012 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-22570488

RESUMO

Serum half-life of IgG is controlled by the neonatal Fc receptor (FcRn) that interacts with the IgG Fc region and may be increased or decreased as a function of altered FcRn binding. Preclinical evaluations of modified IgGs are frequently carried out in mice, but such IgGs may bind differently to mouse and human FcRn (mFcRn and hFcRn). Here, we report a detailed characterization of a matched set of mouse-human chimeric T84.66 scFv-Fc variants with specificity for the tumor carcinoembryonic antigen and mutations in the FcRn-binding site. Binding to soluble mFcRn and hFcRn was measured using in vitro assays, and the results were compared with blood clearance in vivo in normal (mFcRn bearing) and hFcRn transgenic mice. All variants bound better to mFcRn than to hFcRn. The loss of affinity varied among the mutants, however, and also the hierarchy of binding differed depending on the receptor. The mutations had no major impact on binding to the classical Fcγ receptors. Importantly, the trend of blood clearance in both strains of mice correlated with the hierarchy of binding obtained using soluble FcRn. Consequently, in vitro interaction analysis of engineered IgGs regarding their cross-species FcRn binding ability provides information for prediction of in vivo pharmacokinetics.


Assuntos
Antígeno Carcinoembrionário/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Imunoglobulina G/imunologia , Engenharia de Proteínas/métodos , Receptores Fc/imunologia , Receptores de IgG/imunologia , Anticorpos de Cadeia Única/imunologia , Animais , Especificidade de Anticorpos , Reações Cruzadas/imunologia , Humanos , Concentração de Íons de Hidrogênio , Imunoglobulina G/sangue , Radioisótopos do Iodo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Anticorpos de Cadeia Única/farmacocinética , Especificidade da Espécie
3.
Mult Scler J Exp Transl Clin ; 5(2): 2055217319856903, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31223483

RESUMO

BACKGROUND: Multiple sclerosis-associated genetic variants indicate that the adaptive immune system plays an important role in the risk of developing multiple sclerosis. It is currently not well understood how these multiple sclerosis-associated genetic variants contribute to multiple sclerosis risk. CD4+ T cells are suggested to be involved in multiple sclerosis disease processes. OBJECTIVE: We aim to identify CD4+ T cell differential gene expression between multiple sclerosis patients and healthy controls in order to understand better the role of these cells in multiple sclerosis. METHODS: We applied RNA sequencing on CD4+ T cells from multiple sclerosis patients and healthy controls. RESULTS: We did not identify significantly differentially expressed genes in CD4+ T cells from multiple sclerosis patients. Furthermore, pathway analyses did not identify enrichment for specific pathways in multiple sclerosis. When we investigated genes near multiple sclerosis-associated genetic variants, we did not observe significant enrichment of differentially expressed genes. CONCLUSION: We conclude that CD4+ T cells from multiple sclerosis patients do not show significant differential gene expression. Therefore, gene expression studies of all circulating CD4+ T cells may not result in viable biomarkers. Gene expression studies of more specific subsets of CD4+ T cells remain justified to understand better which CD4+ T cell subsets contribute to multiple sclerosis pathology.

4.
PLoS One ; 10(7): e0132957, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26203907

RESUMO

For multiple sclerosis, genome wide association studies and follow up studies have identified susceptibility single nucleotide polymorphisms located in or near CLEC16A at chromosome 16p13.13, encompassing among others CIITA, DEXI and SOCS1 in addition to CLEC16A. These genetic variants are located in intronic or intergenic regions and display strong linkage disequilibrium with each other, complicating the understanding of their functional contribution and the identification of the direct causal variant(s). Previous studies have shown that multiple sclerosis-associated risk variants in CLEC16A act as expression quantitative trait loci for CLEC16A itself in human pancreatic ß-cells, for DEXI and SOCS1 in thymic tissue samples, and for DEXI in monocytes and lymphoblastoid cell lines. Since T cells are major players in multiple sclerosis pathogenesis, we have performed expression analyses of the CIITA-DEXI-CLEC16A-SOCS1 gene cluster in CD4+ and CD8+ T cells isolated from multiple sclerosis patients and healthy controls. We observed a higher expression of SOCS1 and CLEC16A in CD4+ T cells in samples homozygous for the risk allele of CLEC16A rs12927355. Pair-wise linear regression analysis revealed high correlation in gene expression in peripheral T cells of CIITA, DEXI, CLEC16A and SOCS1. Our data imply a possible regulatory role for the multiple sclerosis-associated rs12927355 in CLEC16A.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Lectinas Tipo C/genética , Proteínas de Transporte de Monossacarídeos/genética , Esclerose Múltipla/genética , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Proteínas Supressoras da Sinalização de Citocina/genética , Adulto , Alelos , Estudos de Casos e Controles , Feminino , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Lectinas Tipo C/metabolismo , Pessoa de Meia-Idade , Proteínas de Transporte de Monossacarídeos/metabolismo , Esclerose Múltipla/epidemiologia , Esclerose Múltipla/imunologia , Fatores de Risco , Proteína 1 Supressora da Sinalização de Citocina , Proteínas Supressoras da Sinalização de Citocina/metabolismo , Adulto Jovem
5.
PLoS One ; 10(3): e0117403, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25734800

RESUMO

OBJECTIVE: Determine whether MS-specific DNA methylation profiles can be identified in whole blood or purified immune cells from untreated MS patients. METHODS: Whole blood, CD4+ and CD8+ T cell DNA from 16 female, treatment naïve MS patients and 14 matched controls was profiled using the HumanMethylation450K BeadChip. Genotype data were used to assess genetic homogeneity of our sample and to exclude potential SNP-induced DNA methylation measurement errors. RESULTS: As expected, significant differences between CD4+ T cells, CD8+ T cells and whole blood DNA methylation profiles were observed, regardless of disease status. Strong evidence for hypermethylation of CD8+ T cell, but not CD4+ T cell or whole blood DNA in MS patients compared to controls was observed. Genome-wide significant individual CpG-site DNA methylation differences were not identified. Furthermore, significant differences in gene DNA methylation of 148 established MS-associated risk genes were not observed. CONCLUSION: While genome-wide significant DNA methylation differences were not detected for individual CpG-sites, strong evidence for DNA hypermethylation of CD8+ T cells for MS patients was observed, indicating a role for DNA methylation in MS. Further, our results suggest that large DNA methylation differences for CpG-sites tested here do not contribute to MS susceptibility. In particular, large DNA methylation differences for CpG-sites within 148 established MS candidate genes tested in our study cannot explain missing heritability. Larger studies of homogenous MS patients and matched controls are warranted to further elucidate the impact of CD8+ T cell and more subtle DNA methylation changes in MS development and pathogenesis.


Assuntos
Linfócitos T CD8-Positivos/metabolismo , Metilação de DNA , Genoma Humano , Esclerose Múltipla/genética , Adolescente , Adulto , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Estudos de Casos e Controles , Ilhas de CpG , Feminino , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Modelos Lineares , Pessoa de Meia-Idade , Esclerose Múltipla/patologia , Polimorfismo de Nucleotídeo Único , Análise de Componente Principal , Fatores de Risco , Adulto Jovem
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