DNA vaccination encoding glutamic acid decarboxylase can enhance insulitis and diabetes in correlation with a specific Th2/3 CD4 T cell response in non-obese diabetic mice

CD8+ T cells play an important role in the initiation of insulitis and in the destructive stage leading to insulin-dependent diabetes mellitus. A string of recent studies has led to the identification of numerous HLA-A2-restricted epitopes derived from pancreatic β cell Ags. It is hoped that assays detecting responses of patient PBMC to such epitopes might be instrumental for early diagnosis of β cell-directed autoimmunity and for monitoring trials of immunointervention. However, it remains unclear whether the results of assays studying PBMC reflect responses of islet-infiltrating lymphocytes, and to what extent they correlate with disease risk and/or activity. We have used female and male humanized NOD mice expressing HLA-A2 in addition to murine MHC class I molecules to study spontaneous responses of islet-infiltrating blood, spleen, and lymph node lymphocytes of various age groups to a panel of 16 epitopes. Twelve of these are restricted by HLA-A2, have previously been shown to be recognized by patient CTL, and have identical sequences in human and murine autoantigens. Using an IFN-γ ELISPOT assay, we find highly similar hierarchies of epitope immunodominance in the different T cell compartments, including peripheral blood and pancreatic islets. Moreover, we demonstrate that most of the epitopes eliciting dominant responses in humans display similar status in the mouse model. These results emphasize the potential of humanized mice as tools for studying spontaneous autoimmune CTL responses, and they provide a strong rationale for the development and use of assays monitoring responses of CD8+ PBMC in human type 1 diabetes.

Section: Nsulin-dependent Diabetes Mellitus (Iddm)mentioning

confidence: 99%

Equivalent Specificity of Peripheral Blood and Islet-Infiltrating CD8+ T Lymphocytes in Spontaneously Diabetic HLA-A2 Transgenic NOD Mice

Énée

Martinuzzi

Blancou

et al. 2008

“…pc-GAD were previously described (40). The catalytic domain of IA-2 was amplified by PCR (TaqDNA polymerase; Promega) from ICA512.bdc pCRII (provided by Dr. C. Levy-Marchal, Hospital Robert Debré, Paris, France) using a sense oligonucleotide containing an EcoRI site (5Ј-GGAATTCTAACAT GGACATCTCCACGGG-3Ј; MWG) and an antisense primer containing an XbaI site (5Ј-CGGGCCCTCTAGAGCCTGGGGCAGGGCCTT-3Ј).…”

Section: Plasmid Construction and Mouse Immunizationsmentioning

confidence: 99%

“…Recombinant proteins produced by transiently transfected mammalian cells (293T cells) were detected by immunoblotting. HHDII mice were randomly allocated and DNA vaccinated twice at 6-wk intervals as previously described (40). …”

Section: Plasmid Construction and Mouse Immunizationsmentioning

confidence: 99%

Immunization of HLA Class I Transgenic Mice Identifies Autoantigenic Epitopes Eliciting Dominant Responses in Type 1 Diabetes Patients

Blancou

Mallone²,

Martinuzzi³

et al. 2007

Type 1 diabetes (T1D) results from the autoimmune destruction of pancreatic β cells. CD8+ T cells have recently been assigned a major role in β cell injury. Consequently, the identification of autoreactive CD8+ T cells in humans remains essential for development of therapeutic strategies and of assays to identify aggressive cells. However, this identification is laborious and limited by quantities of human blood samples available. We propose a rapid and reliable method to identify autoantigen-derived epitopes recognized by human CD8+ T lymphocytes in T1D patients. Human histocompatibility leukocyte Ags-A*0201 (HLA-A*0201) transgenic mice were immunized with plasmids encoding the T1D-associated autoantigens: 65 kDa glutamic acid decarboxylase (GAD) or insulinoma-associated protein 2 (IA-2). Candidate epitopes for T1D were selected from peptide libraries by testing the CD8+ reactivity of vaccinated mice. All of the nine-candidate epitopes (five for GAD and four for IA-2) identified by our experimental approach were specifically recognized by CD8+ T cells from newly diagnosed T1D patients (n = 19) but not from CD8+ T cells of healthy controls (n = 20). Among these, GAD114–123, GAD536–545 and IA-2805–813 were recognized by 53%, 25%, and 42% of T1D patients, respectively.

“…To date, a series of different DNA vaccine strategies have been used in the NOD mice to induce Agspecific tolerance, but with variable outcomes. Routes of administration (i.e., intradermal, i.m., or oral), choice of Ag, as well as coadministration of Ag-coding and IL-coding plasmids determine protection versus acceleration of T1D in this model (8)(9)(10)(11)(12)(13). In most cases, these experiments have been carried out without a detailed analysis of the Ag-specific T cell response due to the lack of suitable reagents.…”

mentioning

confidence: 99%

Targeting of a T Cell Agonist Peptide to Lysosomes by DNA Vaccination Induces Tolerance in the Nonobese Diabetic Mouse

Rivas

Driver

Garabatos

et al. 2011

CD4 T cells are crucial effectors in the pathology of type 1 diabetes (T1D). Successful therapeutic interventions for prevention and cure of T1D in humans are still elusive. Recent research efforts have focused on the manipulation of T cells by treatment with DNA. In this paper, we studied the effects of a DNA treatment strategy designed to target antigenic peptides to the lysosomal compartment on a monospecific T cell population termed 2.5mi+ T cells that shares reactivity with the diabetogenic T cell clone BDC-2.5 in the NOD mouse. MHC class II tetramer analysis showed that repeated administrations were necessary to expand 2.5mi+ T cells in vivo. This expansion was independent of Ag presentation by B cells. A single peptide epitope was sufficient to induce protection against T1D, which was not due to Ag-specific T cell anergy. Typical Th2 cytokines such as IL-10 or IL-4 were undetectable in 2.5mi+ T cells, arguing against a mechanism of immune deviation. Instead, the expanded 2.5mi+ T cell population produced IFN-γ similar to 2.5mi+ T cells from naive mice. Protection against T1D by DNA treatment was completely lost in NOD.CD28−/− mice which are largely deficient of natural regulatory T cells (Treg). Although Ag-specific Foxp3+ Treg did not expand in response to DNA treatment, diabetes onset was delayed in Treg-reconstituted and DNA-treated NOD.SCID mice. These observations provide evidence for a Treg-mediated protective mechanism that is independent of the expansion or de novo generation of Ag-specific Treg.