AXL inhibition suppresses early allograft monocyte-to-macrophage differentiation and prolongs allograft survival
Collin Z. Jordan, … , Edward B. Thorp, Xunrong Luo
Collin Z. Jordan, … , Edward B. Thorp, Xunrong Luo
Published January 23, 2024
Citation Information: JCI Insight. 2024;9(5):e178502. https://doi.org/10.1172/jci.insight.178502.
View: Text | PDF
Research Article Immunology Transplantation

AXL inhibition suppresses early allograft monocyte-to-macrophage differentiation and prolongs allograft survival

Abstract

Innate immune cells are important in the initiation and potentiation of alloimmunity in transplantation. Immediately upon organ anastomosis and reperfusion, recipient monocytes enter the graft from circulation and differentiate to inflammatory macrophages to promote allograft inflammation. However, factors that drive their differentiation to inflammatory macrophages are not understood. Here, we show that the receptor tyrosine kinase AXL was a key driver of early intragraft differentiation of recipient infiltrating monocytes to inflammatory macrophages in the presence of allogeneic stimulation and cell-to-cell contact. In this context, the differentiated inflammatory macrophages were capable of efficient alloantigen presentation and allostimulation of T cells of the indirect pathway. Consequently, early and transient AXL inhibition with the pharmacological inhibitor bemcentinib resulted in a profound reduction of initial allograft inflammation and a significant prolongation of allograft survival in a murine heart transplant model. Our results support further investigation of AXL inhibition as part of an induction regimen for transplantation.

Authors

Collin Z. Jordan, Matthew Tunbridge, Irma Husain, Hiroki Kitai, Miriam E. Dilts, Olivia K. Fay, Koki Abe, Catherine Xiang, Jean Kwun, Tomokazu Souma, Edward B. Thorp, Xunrong Luo

×

Figure 1

MC to iMϕ differentiation in response to allogeneic stimulation is dependent on AXL.

Options: View larger image (or click on image) Download as PowerPoint
MC to iMϕ differentiation in response to allogeneic stimulation is depen...
(A) Experimental design schematic demonstrating the in vitro allogeneic coculture model. Congenic CD45.1+ C57BL6/J (B6) MCs were added into coculture with either CD45.2+ syngeneic (B6) or allogenic (BALB/c) BM-derived macrophages (BMDMs). Allogeneic cultures were conducted with or without the AXL inhibitor bemcentinib. FACS analysis was performed after 4 days. (B) Representative flow cytometry plots validating the phenotype of B6 CD45.1+CD11b+Ly6C+F4/80– MCs after isolation via magnetic bead–mediated negative selection, demonstrating ~95% purity of the intended population (n = 3). (C) Representative flow cytometry plots were obtained 4 days after culture of CD45.1+CD11b+Ly6C+ MCs alone (CT), in coculture with syngeneic BMDM (SYN), and in coculture with allogeneic BMDM without bemcentinib (ALLO) or with bemcentinib (ALLO + BEM). Quantification graph of F4/80hi iMϕ frequency under all 4 experimental conditions (n = 4–10 per group), analyzed by 1-way ANOVA with corrected post hoc t tests. **P < 0.01, ****P < 0.0001. (D) Representative flow cytometry plot confirming iMϕ phenotype via absence of CD11c expression and diminishment of CCR2 expression, each respectively compared with an isotype control antibody stain. Normalized mean fluorescence intensity (MFI) of MHCII and CD80 expression gated on iMϕ (n = 4–10 per group). Analysis by nonparametric 1-way ANOVA with post hoc Dunnett’s tests. *P < 0.05, **P < 0.01. (E) Representative flow cytometry plots obtained on day 4 following culture of MCs alone, and coculture of MCs with syngeneic or allogeneic BMDMs in transwell (plots are representative of n = 3 per group).