Transcriptome profiling reveals that cellular composition contributes to the hyaline-vascular variant in unicentric Castleman disease-associated paraneoplastic pemphigus

BACKGROUND: Unicentric Castleman disease (UCD), a rare lymphoproliferative disorder, is frequently complicated by paraneoplastic pemphigus (PNP), an autoimmune mucocutaneous syndrome with high mortality. The hyaline-vascular (HV) histological subtype predominates in UCD-PNP, yet the mechanisms driving vascular hyalinization and stromal dysregulation remain poorly defined. OBJECTIVES: To elucidate the cellular and molecular pathogenesis of UCD-PNP through integrated transcriptomic and cellular analyses, focusing on stromal-immune crosstalk and mechanisms underlying extracellular matrix (ECM) dysregulation. METHODS: We performed bulk RNA sequencing (RNAseq) of lymph node samples from 33 patients with pathologically confirmed UCD-PNP and single-cell RNAseq (scRNAseq) in 5 of them. Analytical approaches included differential expression, pathway enrichment, cellular deconvolution, developmental trajectory inference, ligand-receptor interaction analysis and spatial validation. The functional consequences of identified interactions were assessed using bulk RNAseq and proteomic analysis. RESULTS: Bulk RNAseq highlighted ECM dysregulation, with significant upregulation of collagen genes in UCD-PNP. scRNAseq of 58 811 cells revealed expansion of endothelial cells (ECs), pericytes and fibroblasts, alongside diminished follicular dendritic cells (FDCs). Cell-cell communication analysis identified ECs as primary contributors to collagen and laminin overproduction via COL4A1-ITGA1/ITGB1 and LAMB1-ITGA6/ITGB1 signalling, directly linking EC activity to perivascular hyalinization. UCD-PNP also featured marked plasmablast expansion, IgG class-switching and memory CD4+ T cells driving B-cell hyperactivity. In addition, ligand-receptor analysis revealed a pivotal interaction between EC-derived COL4A1 and CD44 on B cells. Mechanistically, COL4A1 overexpression in ECs upregulated genes involved in ECM organization and remodelling. Furthermore, proteomics revealed EC-B-cell crosstalk-derived vascular basement membrane accumulation (perlecan, encoded by HSPG2) and proinflammatory cytokine release (CCL4) in functional co-cultures. CONCLUSIONS: UCD-PNP pathogenesis centres on aberrant EC expansion and dysregulation, driving simultaneous vascular hyalinization via excessive ECM deposition. Pathological EC-B-cell interactions directly link basement membrane accumulation to proinflammatory signalling. Targeting this EC-driven stromal-immune crosstalk presents a novel therapeutic strategy for UCD-PNP.