Next Generation Sequencing Based Analysis of the T-Cell Receptor Repertoire Reveals an Increase of Alloreactive T-Cell Clones after Kidney Transplantation
1Department of Medicine III - Division of Nephrology and Dialysis, Medical University of Vienna, Vienna, Austria, 2Bioinformatics Research Group, University of Applied Sciences Upper Austria, Hagenberg im Muehlkreis, Austria
Meeting: 2020 American Transplant Congress
Abstract number: B-371
Keywords: Allorecognition, Rejection, T cell clones, T cells
Session Information
Session Name: Poster Session B: Antigen Presentation / Allorecognition / Dendritic Cells
Session Type: Poster Session
Date: Saturday, May 30, 2020
Session Time: 3:15pm-4:00pm
Presentation Time: 3:30pm-4:00pm
Location: Virtual
*Purpose: The development of the alloreactive T-cell repertoire after kidney transplantation (KTX) is not yet fully understood. The combination of mixed lymphocyte reaction (MLR) and next generation sequencing (NGS) of T-cell receptors (TCR) makes in depth analysis of the alloreactive T-cell repertoire possible. Here, we characterize the alloreactive T-cell repertoire from kidney transplant recipients (KTR) and track changes in their alloreactive repertoire post-transplantation.
*Methods: KTRs and their donors were enrolled into our prospective KTX biobank. Thirty-six ml EDTA-blood was collected at the following time points: pre- and ten days post-KTX and at time of management/indication biopsies. Additionally, 36ml blood from donors was collected. PBMCs were isolated using Ficoll centrifugation and cryopreserved. For this analysis eight KTR donor pairs were retrieved from our biobank.
For each pair an MLR for expanding donor-specific T-cells was performed. MLR, pre-KTX and biopsy samples were FACS sorted to obtain CD4+ and CD8+ T-cells. RNA extracted with Trizol was used to prepare TCR-beta NGS libraries which were sequenced on a NextSeq500. Unique molecular identifier guided consensus assembly, followed by alignment to the TCR-locus and clustering of clonotypes was carried out. The alloreactive fraction of a repertoire was computed by adding up respective frequencies for donor-specific clonotypes.
*Results: The median number of clones sequenced from pre-KTX and biopsy samples were 266,091 and 72,328.5 for CD4+ and CD8+ T-cells, respectively. In the CD4+ T-cell compartment an increase in the alloreactive fraction post-transplant was consistently observed for all samples. While V-segment usage was generally quite similar in all three samples (pre-KTX, biopsy and MLR) from a patient, a comparison of Jensen-Shannon divergence revealed that the V-segment usage in MLR samples was significantly different from the V-segment usage in pre-KTX and biopsy samples.
*Conclusions: The fraction of alloreactive T-cells increases post-transplantation. Further studies are needed to unravel unique qualities of the alloreactive repertoire of KTRs experiencing a T-cell mediated rejection.
To cite this abstract in AMA style:
Aschauer C, Jelencsics K, Hu K, Reindl-Schwaighofer R, Vetter J, Heinzel A, Kainz A, Gualdoni G, Schaller S, Winkler S, Oberbauer R. Next Generation Sequencing Based Analysis of the T-Cell Receptor Repertoire Reveals an Increase of Alloreactive T-Cell Clones after Kidney Transplantation [abstract]. Am J Transplant. 2020; 20 (suppl 3). https://atcmeetingabstracts.com/abstract/next-generation-sequencing-based-analysis-of-the-t-cell-receptor-repertoire-reveals-an-increase-of-alloreactive-t-cell-clones-after-kidney-transplantation/. Accessed May 16, 2025.« Back to 2020 American Transplant Congress