Abstract: FR-PO996
Ex Vivo Allogeneic Expansion of Peripheral Blood Mononuclear Cells (PBMCs) from Stable Kidney Transplant Recipients Differentially Regulates Long Noncoding RNA Expression
Session Information
- Transplantation: Basic
October 25, 2024 | Location: Exhibit Hall, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Transplantation
- 2101 Transplantation: Basic
Authors
- Chandraker, Anil K., University of Massachusetts Chan Medical School, Worcester, Massachusetts, United States
- Schreiber, Brittany Lauren, Brigham and Women's Hospital, Boston, Massachusetts, United States
- Tripathi, Sudipta, University of Massachusetts Chan Medical School, Worcester, Massachusetts, United States
Background
We have successfully expanded human Antigen Specific T Regulatory cell enriched Lines (ASTRLs) from stable kidney transplant recipients. Characterization of this preclinical heterogenous cell therapy product to gain greater insight and to interpret how this cellular heterogeneity can be leveraged particularly in its clinical therapeutic applications is pertinent. The purpose of this study is to understand the cellular heterogeneity of ex vivo expanded ASTRLs using cutting edge single cell RNA sequencing. This will provide information about unexplored/ pathways that play a role in cell-cell interaction in heterogenous cell therapy products.
Methods
Single cell RNA sequencing was performed on frozen samples of pre-expanded PBMCs and a post-expanded ASTRL from 5 stable kidney transplant recipients using the 10X Genomics platform. The gene expression counts and TCR/BCR clonotypes were generated using Cellranger pipelines with the human genome reference (GRCh38). The data were imported and analyzed using Seurat and djvdj in R.
Results
Single cell gene expression profile show ASTRLs are enriched for T and NK cell populations and contain very few B cells and monocytes. UMAP clustering distinguishes the CD4+ T cell compartment in all the five ASTRLs as a distinct T cell subset as expected and very similar to the corresponding PBMC samples. Our data shows a number of long noncoding RNA are differentially expressed both in the CD4 and CD8 compartment of ASTRLs in comparison to the PBMC samples. GATA3-AS1, the lncRNA contiguous to GATA3 and essential for the transcription of IL-5 and IL-13, is upregulated 3-fold in the CD4 compartment of ASTRLs, corroborating our previous observation that ASTRLs exhibit a Th2 type Treg phenotype and produce a significantly high levels of IL-13. LINC00402 known as a regulator of allogeneic response in transplantation is 6-fold downregulated in ASTRLs validating the regulatory nature of ASTRLs, that are donor antigen specific. We also observe multiple lncRNA associated with cholesterol biosynthesis and fatty acid oxidation pathways upregulated >2-fold in ASTRLs in comparison to PBMC.
Conclusion
Examination of the differential expression of lncRNAs in ASTRL shows a bias towards fatty acid metabolism and aTh2 like Treg phenotype.
Funding
- Private Foundation Support