SPON1-positive stellate-like stromal cells are associated with fibrotic remodeling and β-cell injury in type 2 diabetes

BACKGROUNDProgressive β-cell failure is central to type 2 diabetes mellitus (T2DM), but the contribution of the islet stromal microenvironment remains incompletely defined.Pancreatic stellate cells can adopt activated extracellular matrix (ECM)-producing states, yet their heterogeneity and relationship to β-cell injury in human T2DM are unclear.We hypothesized that SPON1 expression marks a disease-associated stromal state linked to fibrotic islet remodeling and β-cell injury. AIMTo characterize SPON1-associated stellate-like stromal states and their association with fibrotic remodeling and β-cell injury in T2DM. METHODSWe integrated 159031 cells from pancreatic islet single-cell RNA-sequencing data from 17 control, 11 prediabetic, and 14 T2DM donors with in silico SPON1 knockout, human pancreatic histology and multiplex immunofluorescence, cell-cell communication analysis, and two complementary spatial transcriptomic datasets.Donor-or samplelevel nonparametric, permutation, correlation, and mixed-effects analyses were used as appropriate. RESULTST2DM was associated with altered stromal-state composition, including increased representation of contractile and ECM-enriched transcriptional states.SPON1 was enriched in ECM-associated stromal states, and SPON1-high cells had higher fibrosis/ECM, stellate-activation, and transforming growth factor-β response scores than donor-matched SPON1-low cells (all false discovery rates = 1.1 × 10 -3 ).Virtual knockout 5 / 62 identified predicted regulatory perturbations involving ECM-related genes.SPON1positive alpha-smooth muscle actin-positive stromal cell abundance correlated with the proportion of terminal deoxynucleotidyl transferase dUTP nick-end labeling-positive β cells (Spearman r = 0.629, P < 0.001).T2DM samples had greater peri-islet fibroblast and collagen-positive fibroblast abundance (both false discovery rate = 0.028), and highdamage β-cell regions were closer to SPON1-high stellate-enriched regions (mean difference = -78.5 μm, permutation P = 0.0316).CONCLUSION SPON1-associated stellate-like stromal states are linked to fibrotic remodeling and β-cell injury in T2DM, without establishing lineage-specific identity or a causal role for SPON1.