Single-Islet Proteomics Maps Pseudo-Temporal Islet Immune Responses and Dysfunction in Stage 1 Type 1 Diabetes

ABSTRACT Progressive β-cell dysfunction precedes the onset of type 1 diabetes (T1D), yet the molecular mechanisms driving early T1D development remain poorly understood. Although single-cell RNA-sequencing has uncovered transcript-level changes in human islet cells, it offers limited insight into the heterogeneity of distinct islet microenvironments. Here, we applied a single-islet proteomics workflow to profile intra-donor islet heterogeneity in three stage 1 T1D cases with matched non-diabetic controls and define in situ protein signatures of pseudo-temporal islet dysfunction. Intra-donor analyses of ∼100 individual islets per donor revealed highly consistent proteomic patterns reflecting pseudo-time progression of islet immune responses and β-cell dysfunction. Several pathways, including extracellular matrix remodeling and mRNA processing, were identified as closely associated with progressive islet immune activation and loss of β-cell function. These findings provide robust proteome-wide evidence of the progression of islet dysfunction, offer a valuable resource for investigating early mechanisms of T1D pathogenesis— including novel candidates for functional studies—and underscore the utility of single-islet spatial proteomics for examining islet heterogeneity in T1D. Competing Interest Statement The authors have declared no competing interest.