in-vitro-resident-memory-cd8-t-cell-differentiation-using-epithelial

<em>In Vitro</em> Resident Memory CD8 T Cell Differentiation Using Epithelial Organoid-T Cell Co-culture System

Barrier mucosal tissues play an important role in the differentiation and function of resident T cells. Among the diverse cell populations within these ...

Brown University

Resident memory T cells (TRMs) play a vital role in regional immune defense. Although laboratory rodents have been extensively used to study fundamental TRM biology, poor isolation efficiency and low cell survival rates have limited the implementation of TRM-focused high-throughput assays. Here, we engineer a murine vaginal epithelial organoid (VEO)-CD8 T cell co-culture system that supports CD8 TRM differentiation. These in-vitro-generated TRMs are phenotypically and transcriptionally similar to in vivo TRMs. Pharmacological and genetic approaches showed that transforming growth factor β (TGF-β) signaling plays a crucial role in their differentiation. The VEOs in our model are susceptible to viral infections and the CD8 T cells are amenable to genetic manipulation, both of which will allow a detailed interrogation of antiviral CD8 T cell biology. Altogether we have established a robust in vitro TRM differentiation system that is scalable and can be subjected to high-throughput assays that will rapidly add to our understanding of TRMs.

Epithelial organoid supports resident memory CD8 T cell differentiation.

Inducing robust resident memory T cell (TRM) establishment in mucosal tissues could enhance immunotherapy and vaccine efficacy. However, the factors influencing TRM formation are not fully understood. We present a protocol to coculture murine effector CD8 T cells with epithelial organoids, which serve as a reductionist model for investigating TRM differentiation in vitro. We describe steps for naive CD8 T cell activation, coculture of T cells with vaginal epithelial organoids (VEOs), and analysis of T cell phenotypes via flow cytometry. For complete details on the use and execution of this protocol, please refer to Ulibarri et al..

Article	Year
Epithelial organoid supports resident memory CD8 T cell differentiation. Cell reports\| PubMed ID: 39153200	2024
Protocol for the coculture of murine vaginal epithelial organoids and T cells to induce resident memory CD8 T cell differentiation. STAR protocols\| PubMed ID: 40884788	2025

Julian C. Ramprashad

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