Tissue Derived Non-Classical Monocyte Derived Host Macrophages Protect Against Murine Intestinal Acute Graft-Versus-Host Disease
Authors: Le Duc Dung , Jordán Garrote Ana-Laura , Maria Ranecky , Ottmüller Katja J , Shaikh Haroon , Qureischi Musga , Scheller Lukas , Steinfatt Tim , Brandl Andreas , Hartweg Julia , Brede Christian , Bäuerlein Carina , Riedel Simone S , Tata Nageswara Rao , Mottok Anja , Einsele Hermann , Vafadarnejad Ehsan , Saliba Antoine Emmanuel , Wagers Amy J. , Garbi Natalio , Lutz Manfred B , Beilhack Andreas 
Published: October 2019

Abstract
Acute graft versus host disease (aGVHD) remains a major complication in patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT), the only curative treatment for many malignant hematologic diseases. After initial priming in secondary lymphoid organs, alloreactive donor T cells efficiently migrate to the intestinal tract, liver and skin. We observed that alloreactive effector T cells infiltrating and attacking the lamina propria of the small and large intestines closely interact with intestinal myeloid cells of host origin. Here we asked whether these intimate interactions regulate alloreactive effector T cell responses and how they impact intestinal aGvHD.
To address these questions, we employed non-invasive bioluminescence imaging, fluorescence and confocal microscopy, clinical and histopathologic scoring, flow cytometry and single cell RNA sequencing in murine models of myeloablative, MHC-mismatched allo-HSCT.
In the intestinal lamina propria, we observed that allogeneic T cells closely interacted with CD11b+CD11c+CD103- radio-resistant host type hematopoietic myeloid antigen presenting cells. Selective depletion of intestinal CD11chi or CX3CR1+CD11chi host cells 3 or 8 days after allo-HSCT accelerated alloreactive T cell infiltration, increased T cell mediated inflammatory cytokine production and exacerbated tissue damage resulting in hyperacute lethal aGvHD. These results suggested a strong immunoregulatory effect of these intestinal host-type myeloid cells. Single cell RNA-Seq analysis and flow cytometry (e.g. MHC II, CD11c, F4/80, CD26, CD64, CCR2, CX3CR1), lineage reporter- and defined knockout mice determined these cells as non-classical monocyte derived macrophages as the development and differentiation of these cells did not depend on Flt3, Zbtb46, and CCR2 but rather on CSF-1R and CX3CR1. Adoptive transfer, bone marrow chimeras and parabiosis experiments revealed that these non-classical monocyte derived macrophages differentiated from non-circulating non-classical monocytic precursors. Finally, PD-L1 expression on these intestinal host non-classical monocyte derived macrophages but not on stroma or other hematopoietic cells regulated alloreactive T cell responses in the intestinal tract.
Based on these findings we postulate that a specialized and persistent subpopulation of host non-classical monocyte derived macrophages can potently suppress alloreactive T cells in the lamina propria of the intestinal tract. Fostering the differentiation and function of these tissue resident cells may represent an attractive therapeutic strategy to prevent intestinal aGvHD.

Disclosures
No relevant conflicts of interest to declare.