Bioengineered Extracellular Vesicles Co-delivering PD-L1 and miR-27a-3p Synergistically Reprogram T cells To Treat Inflammatory Bowel Disease [scRNA-Seq]

Current treatments for inflammatory bowel disease (IBD) primarily rely on systemic immunosuppression, which often exhibits limited efficacy and off-target toxicity. Here we present a bioengineered extracellular vesicle (EV) platform designed for inflammatory bowel-targeted delivery of dual-action therapeutics, enabling precise modulation of T cell responses to enhance IBD treatment efficacy. EVs derived from Wharton's jelly mesenchymal stem cells ( WJ-MSCs) were engineered to display membrane-bound PD-L1 and encapsulate miRNA 27a-3p (miR-27a-3p). The platform leverages WJ-MSCs' intrinsic immunomodulatory properties and CXCR4-mediated recruitment to inflamed tissues. PD-L1 not only promotes immunosuppression of effector T cells, but also may enhance EV retention within inflamed sites. Meanwhile, miR-27a-3p shifts the Th17/Treg balance from a pro-inflammatory Th17 phenotype toward an anti-inflammatory Treg phenotype by targeting prohibitin 1. Bulk RNA sequencing of human CD4? T cells treated with the EVs revealed downregulated of Th1/Th17-associated transcripts. In a humanized mouse model of colitis, EV treatment significantly reduced disease severity, diminished intestinal T cell infiltration, and restored mucosal integrity. Single-cell RNA sequencing demonstrated expansion of Treg cells and contraction of effector memory subsets, indicating durable immune modulation. This engineered EV platform represents a novel therapeutic paradigm that combines inflamed tissue targeting and dual immunomodulatory mechanisms to restore immune tolerance in IBD, addressing key limitations of conventional immunosuppressive approaches. Overall design: RNA sequencing was performed on CD4? T cells from spleens of IBD humanized mouse models (human graft), CD4? T cells from human PBMCs, WJ-MSCs, and WJ-MSC–derived exosomes. The study includes single-cell RNA-seq, bulk mRNA-seq, and small RNA-seq to assess transcriptomic and miRNA profiles in the context of IBD and engineered exosome therapy.