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Comprehensive Profiling of Monocytes During Cellular (TCMR) or Mixed Antibody-Mediated Rejection (ABMR) After Kidney Transplantation

C. Macedo1, E. Bailly1, K. Louis2, X. Gu1, M. Lucas1, A. Zeevi1, P. Randhawa1, A. Bansal3, D. Metes1

1Starzl Transplantation Institute, Pittsburgh, PA, 2INSERM UMR-S976, Paris, France, 3University of Pittsburgh, Pittsburgh, PA

Meeting: 2022 American Transplant Congress

Abstract number: 408

Keywords: Alloantibodies, FACS analysis, Mononuclear leukocytes, Rejection

Topic: Clinical Science » Kidney » 44 - Kidney Acute Antibody Mediated Rejection

Session Information

Session Name: Kidney Immunosuppression: Desensitization & Acute Antibody Mediated Rejection

Session Type: Rapid Fire Oral Abstract

Date: Tuesday, June 7, 2022

Session Time: 3:30pm-5:00pm

 Presentation Time: 4:50pm-5:00pm

Location: Hynes Ballroom A

*Purpose: Monocytes play an important role in the immune defense by initiating adaptive immunity. They are heterogeneous, plastic, and present different subsets: classical (CD14++CD16–), intermediate (CD14+CD16+), and non-classical (CD14dimCD16++) that may display a macrophage type-1 (M1) (inflammatory), or type-2 (M2) (reparative) polarization and function. A comprehensive analysis of circulating blood monocyte subsets during rejection after kidney transplantation is still lacking Here, we hypothesize that monocytes with distinct inflammatory programs may differently contribute to TCMR or mixed-ABMR after kidney transplantation (KTx).

*Methods: PBMC collected from 34 KTx patients at the time of a TCMR (n=16) or a mixed-ABMR (n=16) biopsy proven diagnosis were analyzed using 22 multi-color panel with spectral flow cytometry. Magnetic bead-sorted monocytes from 5 TCMR and 5 mixed-ABMR patients were processed for single-cell RNA sequencing. In vitro experiments of FACS-sorted monocytes subsets with either IFNγ or pooled serum DSA from highly sensitized patients (to mimic TCMR or ABMR environment) were set up, and after 5 days, cells were analyzed for phenotypic changes while supernatants were tested for cytokine production.

*Results: A significant phenotypic heterogeneity of blood circulating monocytes was detected between groups. TCMR patients displayed CD11b+CCR2+HLA-DR+ unique classical monocyte clusters while mixed-ABMR patients showed an expanded CD86+CX3CR1+CD163+ intermediate monocytes cell cluster. Transcriptional analyses further highlight that circulating monocytes from TCMR patients presented unique clusters with a classical phenotype and M1-type pro-inflammatory polarization (elevated CXCL11, CRISPLD2, and MMP9 gene expression). Conversely, monocytes from mixed-ABMR patients displayed a significantly expanded intermediate cluster with M2-type polarization (elevated gene transcripts such as PTX3, PRDM1, VEGFA) suggestive of wound healing, vascular repair and tissue fibrosis. In addition, in vitro experiments confirmed that IFNγ treatment triggers classical and intermediate subsets to maintain an activated classical (PD-L1/CD86/CCR2) phenotype, with a M1-like function (IL-12p40/IL-23; MCP1; MIP1α; TNFα). Pooled DSA treatment favors classical and intermediate subsets to differentiate into an intermediate M2-like phenotype with elevated CD86/CD163 expression and pro-inflammatory (IL-1b; IL-6; MCP1; MIP1α) but also anti-inflammatory IL-10 cytokine secretion.

*Conclusions: Our data confirm that monocyte subsets with distinct phenotypes and transcriptional profiles may contribute differently to TCMR or mixed-ABMR. These results suggest a potential need for distinct therapeutic targeting of monocytes during TCMR vs mixed-ABMR rejection.

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To cite this abstract in AMA style:

Macedo C, Bailly E, Louis K, Gu X, Lucas M, Zeevi A, Randhawa P, Bansal A, Metes D. Comprehensive Profiling of Monocytes During Cellular (TCMR) or Mixed Antibody-Mediated Rejection (ABMR) After Kidney Transplantation [abstract]. Am J Transplant. 2022; 22 (suppl 3). https://atcmeetingabstracts.com/abstract/comprehensive-profiling-of-monocytes-during-cellular-tcmr-or-mixed-antibody-mediated-rejection-abmr-after-kidney-transplantation/. Accessed May 9, 2025.

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