Korean J Transplant.  2022 Nov;36(Supple 1):S60. 10.4285/ATW2022.F-1775.

Discovery of cellular and molecular pathways involved in the development of anti-human leukocyte antigen antibody through single cell RNA sequencing in highly sensitized mouse model

• Lee H
• Eum SH
• Yang CW
• Chung BH

• KMID: 2535264
• DOI: http://doi.org/10.4285/ATW2022.F-1775

Abstract

Background
Presence of allo-antibody to human leukocyte antigen (HLA), so called sensitization is an important obstacle for successful kidney transplantation. It is well known that B-cell lineage including antibody producing plasma cells have a major role for the induction of sensitization. However, the specific molecular pathway involved in sensitization has not been fully inves-tigated yet. In this regard, we proposed to observe the specific pathway involved in the sensitization to HLA using allosensitized mouse model using HLA.A2 transgenic mice.
Methods
Wild-type C57BL/6 mice were sensitized with two times of skin allografts from C57BL/6-Tg (HLA-A2.1)1Enge/J mice. We performed single-cell RNA sequencing analysis using splenocytes isolated from allogenic mice (C57BL/6-Tg [HLAA2.1]1Enge/J to C57BL/6) and syngenic control (C57BL/6 to C57BL/6) and compared the gene expression in single cell level to characterize the HLA sensitization.
Results
We generated 10,705 and 17,411 single-cell transcriptomes from allogenic and syngenic control mouse, respectively. Five major cell types (B-cells, T-cells, NK cells, macrophages, and neutrophils) and their transcriptome data were annotated ac-cording to the representative differentially expressed genes (DEGs) of each cell cluster. The percentage of B-cells and T-cells were significantly increased in allogenic mouse, while that of NK cells, macrophages, and neutrophils were decreased. Hsp90aa1 and genes encoding histocompatibility antigen such as H2-Eb1, H2-Ab1, H2-Aa, H2-Oa, H2-DMa, H2-Ob, H2-Q4 were upregulated in B-cells. In addition, GO and KEGG enrichment analyses indicated that the upregulated genes in B-cells were mainly enriched in antigen processing and presentation pathways.
Conclusions
This study identified the comprehensive profiles of complex immune response after transplantation using sin-gle-cell RNA sequencing analysis. The results indicated that overexpressed genes in B-cells after allosensitization were mainly involved in antigen processing and presentation pathways. It may offer detailed understanding of pathogenesis of HLA sensitiza-tion after transplantation and may have implications for the identification of potential therapeutic targets for desensitization.