Objective
Our study aims to untangle how antigen-presenting cells (APCs) and CD8 T cells interact in type 1 diabetes (T1D) to either protect or destroy insulin-producing β cells. We will use a novel cell-labeling technology called uLIPSTIC to trace and identify which APCs directly engage CD8 T cells in pancreatic lymph nodes and pancreas. In a mouse model of T1D, we will analyze the signals exchanged during these encounters to understand how APCs steer CD8 T cells toward tolerance, leaving β cells unharmed, or trigger destructive autoimmunity, leading to β cell destruction. By analyzing APC-CD8 T cell “conversations” before and after T1D develops, we will identify the different APC types contacted by CD8 T cells, their molecular profiles, and how they instruct CD8 T cell behavior. We will also apply this approach to living pancreas tissue slices, allowing us to manipulate and capture changes in CD8 T cell interactions with APCs or β cells directly. Together, these studies will uncover signaling pathways that could be targeted to prevent or stop T1D before β cell damage occurs, offering a roadmap for more precise prevention and intervention strategies in those at risk.
Background Rationale
How type 1 diabetes (T1D) begins and why the immune system shifts from tolerance to β cell destruction is still not fully understood. CD8 T cells are considered key drivers of this process, yet immunotherapies that broadly block their activation only delay, but cannot stop T1D development. To design better prevention and treatment strategies, a deeper knowledge of the upstream mechanisms that keep CD8 T cells protective (“tolerant”) or those that push them to attack β cells is urgently needed. CD8 T cells receive these instructions from another cell type called antigen-presenting cells (APCs). Studying the rare contacts between APCs and CD8 T cells in T1D has been technically very challenging. As a result, the specific APCs involved, their molecular profiles, and how they direct CD8 T cells toward β cell destruction are still unknown. Current technologies either cannot isolate the interacting partners, or they lack the resolution to identify the few cells that actually engaged among the pool of irrelevant “bystanders”. To overcome these limitations, we will use uLIPSTIC, a novel system that tags APCs and CD8 T cells when they “communicate”. Isolating and profiling only these labeled partners provides unprecedented insight into their identities and how they instruct CD8 T cell behavior in T1D. We will combine uLIPSTIC with a well-established T1D mouse model to determine which APCs interact with CD8 T cells and how these contacts drive tolerance or β cell attack. We will also adapt this system to living pancreatic tissue slices, creating a platform to manipulate and investigate interactions between CD8 T cells, APCs, and β cells. These experiments will reveal pathways that could be explored further to prevent or halt T1D before irreversible β cell loss happens.
Description of Project
Type 1 diabetes (T1D) develops when the immune system mistakenly destroys the insulin-producing β cells in the pancreas. This attack is carried out by CD8 T cells, which normally protect the body by eliminating infected or cancerous cells. Notably, similar numbers of CD8 T cells capable of attacking β cells can be detected in the blood of both people with and without T1D. In healthy individuals, their presence alone does not cause disease, indicating that the upstream information CD8 T cells receive from other immune cells determines whether they remain harmless (“tolerant”) or become destructive. Understanding how and where these instructions are delivered is critical for preventing T1D before irreversible β cell loss occurs. The cells that provide these signals to CD8 T cells are called antigen-presenting cells (APCs). APCs constantly show β cell protein fragments to CD8 T cells and guide their responses. Depending on APC type and activation status, some promote tolerance and restrain CD8 T cells, while others trigger immune responses that lead to β cell destruction. Identifying the specific APCs involved and studying their rare, short-lived encounters with CD8 T cells in T1D has been technically very challenging. Standard imaging methods show locations but cannot isolate interacting partners for detailed analyses, while advanced single-cell RNA sequencing mostly captures surrounding “bystanders” rather than the few cells that actually engaged. To overcome these limitations, we will use uLIPSTIC (universal Labeling Immune Partnerships by SorTagging Intercellular Contacts), a novel technology that specifically tags partner cells after they “communicate” in vivo. In this system, when a CD8 T cell contacts an APC, a label is transferred between them, like a molecular “lipstick mark from a kiss”, allowing isolation and analysis of only the cells that actually interacted. Using uLIPSTIC in a well-established mouse model of T1D, we will map the APCs and any β cells that engage CD8 T cells under conditions that either promote tolerance or trigger autoimmune attack. Our project will pursue three main objectives: (1) identify and characterize the APC types that contact CD8 T cells in pancreatic lymph nodes and pancreas; (2) determine how these APCs influence CD8 T cell behavior, revealing which promote tolerance and which induce destructive responses, along with the molecular pathways involved; and (3) apply this assay to a living pancreatic tissue slice system to manipulate and capture CD8 T cell interactions with APCs or β cells directly, providing a foundation for future human tissue studies. By generating the first interaction-resolved map of APC-CD8 T cell crosstalk in T1D, we aim to pinpoint the earliest checkpoints where immune tolerance fails. This knowledge will guide the development of immunotherapies that selectively block harmful APC-CD8 T cell encounters while preserving protective ones, offering the potential to prevent β cell loss. Ultimately, we seek to understand why CD8 T cells capable of attacking β cells remain harmless in some people but turn destructive in others, informing precise strategies to prevent and treat T1D.
Anticipated Outcome
Our experiments will uncover which signaling pathways become activated when antigen-presenting cells (APCs) interact with CD8 T cells and how these mechanisms determine whether immune tolerance is maintained, leaving β cells unharmed, or whether β cell-destructive autoimmunity develops. Using uLIPSTIC, we will define APC signature profiles linked to each outcome and identify potential targets for precise immunotherapies or preventive interventions. Adapting this assay to a living pancreatic slice platform will allow manipulation, such as drug testing, and detailed study of APC- or β cell-CD8 T cell interactions in T1D. These experiments will also lay the foundation for future application to human tissues.
Relevance to T1D
A better understanding of how antigen-presenting cells (APCs) interact with CD8 T cells in T1D is urgently needed to develop therapies that prevent or halt disease progression. Current technologies, such as traditional microscopy or single-cell RNA sequencing, are not capable of directly profiling the few cells that actually interact in T1D. Using uLIPSTIC, we can, for the first time, recover and comprehensively analyze APCs and their CD8 T cell partners after confirmed cell-cell contacts. This will reveal the cellular and molecular mechanisms that drive T1D and identify signaling pathways that can be targeted to design new or improved therapies aimed at preserving β cells and preventing disease progression.