Coxsackievirus infection induces direct pancreatic ß cell killing but poor antiviral CD8+ T cell responses.

Coxsackievirus B (CVB) infection of pancreatic ß cells is associated with ß cell autoimmunity and type 1 diabetes. We investigated how CVB affects human ß cells and anti-CVB T cell responses. ß cells were efficiently infected by CVB in vitro, down-regulated human leukocyte antigen (HLA) class I, and presented few, selected HLA-bound viral peptides. Circulating CD8+ T cells from CVB-seropositive individuals recognized a fraction of these peptides; only another subfraction was targeted by effector/memory T cells that expressed exhaustion marker PD-1. T cells recognizing a CVB epitope cross-reacted with ß cell antigen GAD. Infected ß cells, which formed filopodia to propagate infection, were more efficiently killed by CVB than by CVB-reactive T cells. Our in vitro and ex vivo data highlight limited CD8+ T cell responses to CVB, supporting the rationale for CVB vaccination trials for type 1 diabetes prevention. CD8+ T cells recognizing structural and nonstructural CVB epitopes provide biomarkers to differentially follow response to infection and vaccination.

Coxsackievirus infection induces direct pancreatic ß-cell killing but poor anti-viral CD8+ T-cell responses.

Coxsackievirus B (CVB) infection of pancreatic ß cells is associated with ß-cell autoimmunity. We investigated how CVB impacts human ß cells and anti-CVB T-cell responses. ß cells were efficiently infected by CVB in vitro, downregulated HLA Class I and presented few, selected HLA-bound viral peptides. Circulating CD8+ T cells from CVB-seropositive individuals recognized only a fraction of these peptides, and only another sub-fraction was targeted by effector/memory T cells that expressed the exhaustion marker PD-1. T cells recognizing a CVB epitope cross-reacted with the ß-cell antigen GAD. Infected ß cells, which formed filopodia to propagate infection, were more efficiently killed by CVB than by CVB-reactive T cells. Thus, our in-vitro and ex-vivo data highlight limited T-cell responses to CVB, supporting the rationale for CVB vaccination trials for type 1 diabetes prevention. CD8+ T cells recognizing structural and non-structural CVB epitopes provide biomarkers to differentially follow response to infection and vaccination.

Pancreas Whole Tissue Transcriptomics Highlights the Role of the Exocrine Pancreas in Patients With Recently Diagnosed Type 1 Diabetes.

Although type 1 diabetes (T1D) is primarily a disease of the pancreatic beta-cells, understanding of the disease-associated alterations in the whole pancreas could be important for the improved treatment or the prevention of the disease. We have characterized the whole-pancreas gene expression of patients with recently diagnosed T1D from the Diabetes Virus Detection (DiViD) study and non-diabetic controls. Furthermore, another parallel dataset of the whole pancreas and an additional dataset from the laser-captured pancreatic islets of the DiViD patients and non-diabetic organ donors were analyzed together with the original dataset to confirm the results and to get further insights into the potential disease-associated differences between the exocrine and the endocrine pancreas. First, higher expression of the core acinar cell genes, encoding for digestive enzymes, was detected in the whole pancreas of the DiViD patients when compared to non-diabetic controls. Second, In the pancreatic islets, upregulation of immune and inflammation related genes was observed in the DiViD patients when compared to non-diabetic controls, in line with earlier publications, while an opposite trend was observed for several immune and inflammation related genes at the whole pancreas tissue level. Third, strong downregulation of the regenerating gene family (REG) genes, linked to pancreatic islet growth and regeneration, was observed in the exocrine acinar cell dominated whole-pancreas data of the DiViD patients when compared with the non-diabetic controls. Fourth, analysis of unique features in the transcriptomes of each DiViD patient compared with the other DiViD patients, revealed elevated expression of central antiviral immune response genes in the whole-pancreas samples, but not in the pancreatic islets, of one DiViD patient. This difference in the extent of antiviral gene expression suggests different statuses of infection in the pancreas at the time of sampling between the DiViD patients, who were all enterovirus VP1+ in the islets by immunohistochemistry based on earlier studies. The observed features, indicating differences in the function, status and interplay between the exocrine and the endocrine pancreas of recent onset T1D patients, highlight the importance of studying both compartments for better understanding of the molecular mechanisms of T1D.

Genetic and Environmental Interaction in Type 1 Diabetes: a Relationship Between Genetic Risk Alleles and Molecular Traits of Enterovirus Infection?

PURPOSE OF REVIEW: We provide an overview of the current knowledge regarding the natural history of human type 1 diabetes (T1D) and the documented associations between virus infections (in particular the enteroviruses) and disease development. We review studies that examine whether T1D-specific risk alleles in genes involved in the function of the immune system can alter susceptibility to virus infections or affect the magnitude of the host antiviral response. We also highlight where the major gaps in our knowledge exist and consider possible implications that new insights gained from the discussed gene-environment interaction studies may bring. RECENT FINDINGS: A commonality between several of the studied T1D risk variants studied is their role in modulating the host immune response to viral infection. Generally, little support exists indicating that the risk variants increase susceptibility to infection and moreover, they usually appear to predispose the immune system towards a hyper-reactive state, decrease the risk of infection, and/or favor the establishment of viral persistence. In conclusion, although the current number of studies is limited, this type of research can provide important insights into the mechanisms that are central to disease pathogenesis and further describe how genetic and environmental factors jointly influence the risk of T1D development. The latter may provide genetic markers that could be used for patient stratification and for the selection of method(s) for disease prevention.

Early Detection of Peripheral Blood Cell Signature in Children Developing ß-Cell Autoimmunity at a Young Age.

The appearance of type 1 diabetes (T1D)-associated autoantibodies is the first and only measurable parameter to predict progression toward T1D in genetically susceptible individuals. However, autoantibodies indicate an active autoimmune reaction, wherein the immune tolerance is already broken. Therefore, there is a clear and urgent need for new biomarkers that predict the onset of the autoimmune reaction preceding autoantibody positivity or reflect progressive ß-cell destruction. Here we report the mRNA sequencing-based analysis of 306 samples including fractionated samples of CD4+ and CD8+ T cells as well as CD4-CD8- cell fractions and unfractionated peripheral blood mononuclear cell samples longitudinally collected from seven children who developed ß-cell autoimmunity (case subjects) at a young age and matched control subjects. We identified transcripts, including interleukin 32 (IL32), that were upregulated before T1D-associated autoantibodies appeared. Single-cell RNA sequencing studies revealed that high IL32 in case samples was contributed mainly by activated T cells and NK cells. Further, we showed that IL32 expression can be induced by a virus and cytokines in pancreatic islets and ß-cells, respectively. The results provide a basis for early detection of aberrations in the immune system function before T1D and suggest a potential role for IL32 in the pathogenesis of T1D.

Genome-wide Analysis of STAT3-Mediated Transcription during Early Human Th17 Cell Differentiation.

The development of therapeutic strategies to combat immune-associated diseases requires the molecular mechanisms of human Th17 cell differentiation to be fully identified and understood. To investigate transcriptional control of Th17 cell differentiation, we used primary human CD4+ T cells in small interfering RNA (siRNA)-mediated gene silencing and chromatin immunoprecipitation followed by massive parallel sequencing (ChIP-seq) to identify both the early direct and indirect targets of STAT3. The integrated dataset presented in this study confirms that STAT3 is critical for transcriptional regulation of early human Th17 cell differentiation. Additionally, we found that a number of SNPs from loci associated with immune-mediated disorders were located at sites where STAT3 binds to induce Th17 cell specification. Importantly, introduction of such SNPs alters STAT3 binding in DNA affinity precipitation assays. Overall, our study provides important insights for modulating Th17-mediated pathogenic immune responses in humans.

Comparative analysis of human and mouse transcriptomes of Th17 cell priming.

Uncontrolled Th17 cell activity is associated with cancer and autoimmune and inflammatory diseases. To validate the potential relevance of mouse models of targeting the Th17 pathway in human diseases we used RNA sequencing to compare the expression of coding and non-coding transcripts during the priming of Th17 cell differentiation in both human and mouse. In addition to already known targets, several transcripts not previously linked to Th17 cell polarization were found in both species. Moreover, a considerable number of human-specific long non-coding RNAs were identified that responded to cytokines stimulating Th17 cell differentiation. We integrated our transcriptomics data with known disease-associated polymorphisms and show that conserved regulation pinpoints genes that are relevant to Th17 cell-mediated human diseases and that can be modelled in mouse. Substantial differences observed in non-coding transcriptomes between the two species as well as increased overlap between Th17 cell-specific gene expression and disease-associated polymorphisms underline the need of parallel analysis of human and mouse models. Comprehensive analysis of genes regulated during Th17 cell priming and their classification to conserved and non-conserved between human and mouse facilitates translational research, pointing out which candidate targets identified in human are worth studying by using in vivo mouse models.

Gut-adipose tissue axis in hepatic fat accumulation in humans.

BACKGROUND & AIMS: Recent evidence suggests that in animals gut microbiota composition (GMC) affects the onset and progression of hepatic fat accumulation. The aim of this study was to investigate in humans whether subjects with high hepatic fat content (HHFC) differ in their GMC from those with low hepatic fat content (LHFC), and whether these differences are associated with body composition, biomarkers and abdominal adipose tissue inflammation. METHODS: Hepatic fat content (HFC) was measured using proton magnetic resonance spectroscopy ((1)H MRS). Fecal GMC was profiled by 16S rRNA fluorescence in situ hybridization and flow cytometry. Adipose tissue gene expression was analyzed using Affymetrix microarrays and quantitative PCR. RESULTS: The HHFC group had unfavorable GMC described by lower amount of Faecalibacterium prausnitzii (FPrau) (p<0.05) and relatively higher Enterobacteria than the LHFC group. Metabolically dysbiotic GMC associated with HOMA-IR and triglycerides (p<0.05 for both). Several inflammation-related adipose tissue genes were differentially expressed and correlated with HFC (p<0.05). In addition, the expression of certain genes correlated with GMC dysbiosis, i.e., low FPrau-to-Bacteroides ratio. CONCLUSIONS: HHFC subjects differ unfavorably in their GMC from LHFC subjects. Adipose tissue inflammation may be an important link between GMC, metabolic disturbances, and hepatic fat accumulation.

Early T helper cell programming of gene expression in human.

Molecular mechanisms guiding naïve T helper cell differentiation into functionally specified effector cells are intensively studied. The rapidly growing knowledge is mainly achieved by using mouse cells or disease models. Comparatively exiguous data is gathered from human primary cells although they provide the "ultimate model" for immunology in man, have been exploited in many original studies paving the way for the field, and can be analyzed more easily than ever with the help of modern technology and methods. As usage of mouse models is unavoidable in translational research, parallel human and mouse studies should be performed to assure the relevancy of the hypothesis created during the basic research. In this review, we give an overview on the status of the studies conducted with human primary cells aiming at elucidating the mechanisms instructing the priming of T helper cell subtypes. The special emphasis is given to the recent high-throughput studies. In addition, by comparing the human and mouse studies we intend to point out the regulatory mechanisms and questions which are lacking examination with human primary cells.

Gene expression profiling of immune-competent human cells exposed to engineered zinc oxide or titanium dioxide nanoparticles.

A comprehensive in vitro assessment of two commercial metal oxide nanoparticles, TiO2 and ZnO, was performed using human monocyte-derived macrophages (HMDM), monocyte-derived dendritic cells (MDDC), and Jurkat T cell leukemia-derived cell line. TiO2 nanoparticles were found to be non-toxic whereas ZnO nanoparticles caused dose-dependent cell death. Subsequently, global gene expression profiling was performed to identify transcriptional response underlying the cytotoxicity caused by ZnO nanoparticles. Analysis was done with doses 1 µg/ml and 10 µg/ml after 6 and 24 h of exposure. Interestingly, 2703 genes were significantly differentially expressed in HMDM upon exposure to 10 µg/ml ZnO nanoparticles, while in MDDCs only 12 genes were affected. In Jurkat cells, 980 genes were differentially expressed. It is noteworthy that only the gene expression of metallothioneins was upregulated in all the three cell types and a notable proportion of the genes were regulated in a cell type-specific manner. Gene ontology analysis revealed that the top biological processes disturbed in HMDM and Jurkat cells were regulating cell death and growth. In addition, genes controlling immune system development were affected. Using a panel of modified ZnO nanoparticles, we obtained an additional support that the cellular response to ZnO nanoparticles is largely dependent on particle dissolution and show that the ligand used to modify ZnO nanoparticles modulates Zn(2+) leaching. Overall, the study provides an extensive resource of transcriptional markers for mediating ZnO nanoparticle-induced toxicity for further mechanistic studies, and demonstrates the value of assessing nanoparticle responses through a combined transcriptomics and bioinformatics approach.