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Unique patterns of CD8+ T-cell-mediated organ damage in the Act-mOVA/OT-I model of acute graft-versus-host disease

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Abstract

T-cell receptor (TCR)-transgenic models of acute graft-versus-host disease (aGvHD) offer a straightforward and highly controlled approach to study the mechanisms and consequences of T-cell activation following allogeneic hematopoietic stem cell transplantation (aHSCT). Here, we report that aHSCT involving OT-I mice as donors, carrying an ovalbumin-specific CD8+ TCR, and Act-mOVA mice as recipients, expressing membrane-bound ovalbumin driven by the β-actin promoter, induces lethal aGvHD in a CD8+ T-cell-dependent, highly reproducible manner, within 4–7 days. Tracking of UBC-GFP/OT-I graft CD8+ T cells disclosed heavy infiltration of the gastrointestinal tract, liver, and lungs at the onset of the disease, and histology confirmed hallmark features of gastrointestinal aGVHD, hepatic aGvHD, and aGvHD-associated lymphocytic bronchitis in infiltrated organs. However, T-cell infiltration was virtually absent in the skin, a key target organ of human aGvHD, and histology confirmed the absence of cutaneous aGVHD, as well. We show that the model allows studying CD8+ T-cell responses in situ, as selective recovery of graft CD45.1/OT-I CD8+ T cells from target organs is simple and feasible by automated tissue dissociation and subsequent cell sorting. Assessment of interferon-gamma production by flow cytometry, granzyme-B release by ELISA, TREC assay, and whole-genome gene expression profiling confirmed that isolated graft CD8+ T cells remained intact, underwent clonal expansion, and exerted effector functions in all affected tissues. Taken together, these data demonstrate that the OT-I/Act-mOVA model is suitable to study the CD8+ T-cell-mediated effector mechanisms in a disease closely resembling fatal human gastrointestinal and hepatic aGVHD that may develop after aHSCT using HLA-matched unrelated donors.

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Abbreviations

aGvHD:

Acute graft-versus-host disease

aHSCT:

Allogeneic hematopoietic stem cell transplantation

Act:

Chicken beta-actin promoter

APC:

Antigen-presenting cell

CMV:

Cytomegalovirus

CTL:

Cytotoxic T lymphocyte

DEG:

Differentially expressed gene

ELISA:

Enzyme-linked immunosorbent assay

FDR:

False discovery rate

GFP:

Green fluorescent protein

GI:

Gastrointestinal

GSEA:

Gene set enrichment analysis

HE:

Hematoxylin-eosin staining

HUGO:

Human Genome Organisation

IHC:

Immunohistochemistry

K14:

Cytokeratin 14 promoter

miHA:

Minor histocompatibility antigen

mOVA:

Membrane-bound chicken ovalbumin

MUD:

MHC-matched unrelated donor

NES:

Normalized enrichment score

PBMC:

Peripheral blood mononuclear cell

PDGF:

Platelet-derived growth factor

sjTREC:

Signal joint T-cell receptor excision circles

sOVA:

Soluble chicken ovalbumin

TBI:

Total body irradiation

UBC:

Human ubiqutin C promoter

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Acknowledgments

The authors would like to thank Tamás Masszi and András Falus for critical reading of the manuscript.

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    Authors and Affiliations

    1. Hungarian Academy of Sciences-Semmelweis University, ”Lendület” Experimental and Translational Immunomics Research Group, 1089, Budapest, Hungary

      Barbara Érsek, Nikolett Lupsa, Anett Tóth, Andor Horváth, Bence Bagita, András Bencsik & Zoltán Pós

    2. Department of Genetics, Cell and Immunobiology, Semmelweis University, 1089, Budapest, Hungary

      Barbara Érsek, Nikolett Lupsa, Anett Tóth, Andor Horváth, Viktor Molnár, Bence Bagita, András Bencsik, Edit I. Buzás & Zoltán Pós

    3. 1st Department of Pathology and Experimental Cancer Research, Semmelweis University, 1085, Budapest, Hungary

      Péter Pócza & András Matolcsy

    4. Csertex Research Laboratory, 1037, Budapest, Hungary

      Viktor Molnár

    5. “Frédéric Joliot-Curie” Institute for Radiobiology and Radiohygiene, 1221, Budapest, Hungary

      Hargita Hegyesi

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    Correspondence to Zoltán Pós.

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    The authors declare that they have no conflict of interest.

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    This work was supported by the Hungarian Academy of Sciences (“Lendület” LP2012-49/2012), and the Hungarian National Research, Development and Innovation Office (OTKA K 116340).

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    B. Érsek and N. Lupsa contributed equally to this work.

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    18_2016_2237_MOESM1_ESM.tif

    Supplementary Fig. 1: Successful elimination host CD8+ T cells by lethal dose TBI. Grafting CD45.1 bone marrow into CD45.2 hosts following lethal dose TBI confirms successful elimination of host CTLs by Day 7. Full replacement of host CTLs with graft-derived CTLs occurs by aHSCT + Day 14. Representative flow cytometric analysis of the peripheral blood of affected animals (TIFF 1558 kb)

    18_2016_2237_MOESM2_ESM.xlsx

    Supplementary Table 1: Differentially expressed genes discriminating between CD45.1/OT-I CD8+ T cells grafted into Act-mOVA mive before (aHSCT + Day 0) and 4 days after grafting (aHSCT + Day 4). Table of 2514 genes differentially expressed by CD45.1/OT-I CD8+ T cells in the graft (aHSCT + Day 0), and graft-derived T cells infiltrating the small intestine, lung, and liver of Act-mOVA recipients 4 days later (aHSCT + Day 4). Agilent microarray probeset IDs, HUGO Gene Symbols, uncorrected, and Benjamini--Hochberg corrected results of One-Way ANOVA followed by Tukey’s all pairwise analysis, p values, fold-changes, gene names, full gene annotations, and Agilent probeset descriptions are shown (XLSX 867 kb)

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    Érsek, B., Lupsa, N., Pócza, P. et al. Unique patterns of CD8+ T-cell-mediated organ damage in the Act-mOVA/OT-I model of acute graft-versus-host disease. Cell. Mol. Life Sci. 73, 3935–3947 (2016). https://doi.org/10.1007/s00018-016-2237-7

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