Critical Role of Macrophages in Checkpoint Blockade Induced Autoimmune Diabetes in the NOD Mouse

Abstract We confirmed that the PD-1/PD-L1 regulatory pathway is one control mechanism of autoimmune diabetes in NOD mice: administration of anti-PD-1 but not anti-CTLA-4 antibodies rapidly induced diabetes in young mice showing a control phase established very early in the autoimmune process. We confirmed that both CD4 and CD8 T cells and resident macrophages were essential for the PD-1 regulation to take effect. We performed an unbiased single-cell RNA sequencing analysis of islets after anti-PD-1 treatment. We identified a subpopulation of highly activated macrophage showing an IFNγ signature with high level of CXCL9 and PD-L1 and a monocyte signature distinguishable by the markers Ly6c and CCR2. Flow cytometry analysis of islets upon anti-PD-1 treatment identified a predominance of the monocyte derived macrophages compared to the islet resident macrophages. Depleting blood monocytes by clodronate liposomes inhibited the recruitment of the monocyte derived macrophages and prevented the development of acute diabetes induced by anti-PD-1 treatment. To analyze the pathogenicity of this monocyte-derived macrophages, we performed cytotoxicity assays by co-culturing macrophages stimulated with LPS&IFNγ and a β-cell line Min6: the highly activated macrophage killed β-cells in a manner dependent on IFNγ and inducible nitric oxide. In sum, our study shows highly activated macrophage derived from monocytes are cytotoxic for beta cells and promoting diabetes progression. Our findings also indicate that a consequence of checkpoint blockade is the recruitment and activation of cytocidal macrophages derived from monocytes. Targeting monocyte-derived macrophages may prevent some of the checkpoint blockade related immune-adverse effects.