Background/Purpose: Many rheumatic diseases are driven by chronic, repeated activation of Toll-like receptors (TLR) causing the initiation and perpetuation of disease. However, TLR-activated innate immune cells undergo the well-known immunoregulatory phenomenon of TLR tolerance, whereby the initial activation of TLRs results in impaired proinflammatory responses to subsequent TLR stimuli. Therefore, it is unclear how TLR-driven inflammation persists in vivo. Our well-characterized model of repeated TLR9-driven inflammation demonstrates that repeated activation of TLR9 results in a feed-forward, interleukin (IL)-12-mediated inflammatory response culminating in systemic immunopathology rather than tolerance. Using this model, we dissect the cellular mechanisms utilized to bypass TLR9 tolerance in vivo.

Methods: In vitro mixed TLR9 sufficient and deficient cell cultures were stimulated with CpG1826, a TLR9 agonist, to demonstrate that IL-12 is a readout of cell-intrinsic TLR9 activation. Yet40 IL-12-reporter mice were treated with 4 doses of CpG1826 before tissue leukocytes were isolated and analyzed by flow cytometry to identify the IL-12-producing cells. Bone marrow, peripheral blood, and tissue leukocytes from CpG-treated mice were stained for myeloid progentor cell markers to determine if myelopoiesis is altered during TLR9-mediated systemic inflammation.

Results: We demonstrate that inflammatory monocyte-derived dendritic cells increase in frequency and total numbers during TLR9-mediated systemic inflammation, and are the key IL-12-producing cell in this model. To determine the origin of these inflammatory tissue-invading cells, we tested peripheral blood and bone marrow of CpG-treated mice to enumerate the number of inflammatory monocytes and myeloid progenitor cells. To our surprise, the total number of peripheral blood inflammatory monocytes and bone marrow myeloid precursors was unchanged during TLR9-induced systemic inflammation. In contrast, extramedullary myeloid precursors were markedly increased in peripherally inflamed tissues of CpG-treated mice.

Conclusion: Our data highlight an important mechanism whereby persistent TLR9 activation bypasses TLR tolerance in vivo by increasing extramedullary myelopoiesis. Increased extramedullary myelopoiesis provides a continuous source of new TLR9-responsive cells that perpetuates TLR-driven inflammation. These new insights into the mechanisms driving persistent TLR-mediated inflammation may lead to novel therapeutic targets to ameliorate TLR9-mediated rheumatic diseases by intervening to suppress TLR-driven extramedullary myelopoiesis.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/extramedullary-myelopoiesis-drives-persistent-toll-like-receptor-mediated-inflammation/