Poster Presentations

Start Date

20-5-2014 12:30 PM

Description

Type 1 diabetes (T1D) is an autoimmune disease characterized by the activation of lymphocytes against insulin-producing β-cells in the pancreas. In humans, CD8+ T cells are predominantly found in sites of insulitis and are considered to be one of the main drivers of β-cell destruction, thus indicating the need to analyze the frequency and function of these autoreactive CD8+ T cells. Peripheral blood mononuclear cells (PBMC) from individuals with long-term T1D were stained ex vivo for T cell surface markers and HLA-A2 pentamers containing known islet-associated epitopes to determine if there are autoreactive CD8+ T cells circulating in the periphery. All T1D donors tested had at least one detectable autoreactive CD8 T cell population and the frequencies of these autoantigen-specific T cells were comparable to previously published data from T1D individuals. We then developed a method of establishing CD8 T cell lines by co-culturing negatively isolated CD8 T cells and peptide-pulsed monocyte-derived dendritic cells from the PBMC of one T1D donor (A*02:01, A*33:01, B*14:02, B*40:01, DRB1*01:02, DRB1*04:04). We expanded a CD8 T cell line specific to the preproinsulin peptide PPI15-24. This cell line produced IFN-γ and expressed CD107a in the presence of PPI15-24-pulsed target cells, but not to an unrelated peptide or media alone. Using a similar approach, we were able to generate CD8 T cell lines from the same T1D donor that were cytotoxic to target cells pulsed with the autoantigens glutamic acid decarboxylase peptide (GAD65114-123) and islet-specific glucose-6-phosphatase catalytic subunit-related protein peptide (IGRP265-273). These autoreactive T cell lines can be utilized in in vivo assays using humanized mouse models to further understand the mechanism of β-cell destruction and disease progression. Studying the functionalities of these autoreactive T cells will also provide insights into identifying immune correlates to better assess both novel and existing immunotherapeutic strategies for T1D.

Comments

Abstract of poster presented at the 2014 UMass Center for Clinical and Translational Science Research Retreat, held on May 20, 2014 at the University of Massachusetts Medical School, Worcester, Mass.

Creative Commons License

Creative Commons Attribution-Noncommercial-Share Alike 3.0 License
This work is licensed under a Creative Commons Attribution-Noncommercial-Share Alike 3.0 License.

 
May 20th, 12:30 PM

Detection of CD8+ T cell Responses in Individuals with Long-term Type 1 Diabetes and Generation of Human CD8+ T Cell Lines Specific to Islet-associated Autoantigens

Type 1 diabetes (T1D) is an autoimmune disease characterized by the activation of lymphocytes against insulin-producing β-cells in the pancreas. In humans, CD8+ T cells are predominantly found in sites of insulitis and are considered to be one of the main drivers of β-cell destruction, thus indicating the need to analyze the frequency and function of these autoreactive CD8+ T cells. Peripheral blood mononuclear cells (PBMC) from individuals with long-term T1D were stained ex vivo for T cell surface markers and HLA-A2 pentamers containing known islet-associated epitopes to determine if there are autoreactive CD8+ T cells circulating in the periphery. All T1D donors tested had at least one detectable autoreactive CD8 T cell population and the frequencies of these autoantigen-specific T cells were comparable to previously published data from T1D individuals. We then developed a method of establishing CD8 T cell lines by co-culturing negatively isolated CD8 T cells and peptide-pulsed monocyte-derived dendritic cells from the PBMC of one T1D donor (A*02:01, A*33:01, B*14:02, B*40:01, DRB1*01:02, DRB1*04:04). We expanded a CD8 T cell line specific to the preproinsulin peptide PPI15-24. This cell line produced IFN-γ and expressed CD107a in the presence of PPI15-24-pulsed target cells, but not to an unrelated peptide or media alone. Using a similar approach, we were able to generate CD8 T cell lines from the same T1D donor that were cytotoxic to target cells pulsed with the autoantigens glutamic acid decarboxylase peptide (GAD65114-123) and islet-specific glucose-6-phosphatase catalytic subunit-related protein peptide (IGRP265-273). These autoreactive T cell lines can be utilized in in vivo assays using humanized mouse models to further understand the mechanism of β-cell destruction and disease progression. Studying the functionalities of these autoreactive T cells will also provide insights into identifying immune correlates to better assess both novel and existing immunotherapeutic strategies for T1D.

 

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