Background/Purpose: Monocytes and macrophages are essential to innate immunity but also propagate the inflammatory response in autoimmune arthritis and lupus nephritis.  Spontaneous development of inflammatory monocytes (Ly6C^hi CCR2⁺) and subsequent differentiation to residential monocytes (Ly6C^lo CX₃CR1^hi) from murine bone marrow precursor cells is an enigmatic process that involves numerous transcription factors.  Understanding the signaling pathways involved in myeloid development could help identify novel treatment targets for inflammatory diseases.  

Methods: We developed a murine myeloid progenitor cell line with CCR2^RFP CX₃CR1^GFP reporters to track monocyte differentiation.  Small molecule library screen was used to identify biological pathways involved in monocyte development.  Activation of mTOR substrates was measured by intracellular flow cytometry.  Key findings were confirmed using pharmacologic inhibition and genetic disruption.

Results: Using a novel progenitor cell line that recapitulates the monocyte development continuum, we identified an essential role of mechanistic target of rapamycin complex 1 (mTORC1) on the generation of inflammatory monocyte through a small molecule library screen.  Basal mTORC1 signaling was more prominent in monocytes than other cell subsets in wild-type mice and humanized mice.  Inhibition of mTORC1 by rapamycin or disruption of Raptor (an essential component of mTORC1) impaired monocyte development, while deletion of TSC-1 (a negative regulator of mTORC1) accelerated the differentiation into mature macrophages.  Correspondingly, Raptor ^fl/fl UBC-cre/ERT2 mice susceptible to tamoxifen-mediated deletion of Raptor demonstrated a rapid, inducibe depletion of circulating inflammatory monocytes, confirm an essential non-redundant role of mTORC1 in monocyte development in vivo. Curiously, Raptor-deficient progenitors in the bone marrow spontaneously differentiated into Ly6C^lo CX₃CR1^low myeloid cells with characteristics of alternatively-activated macrophages.   

Conclusion: Using a combination of in vitro and in vivo approaches, we identified a previously unrecognized role of mTORC1 in the normal development of inflammatory monocytes.  These findings support the potential utility of mTORC1 inhibitors in monocyte- and macrophage-mediated inflammatory diseases.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/mtor-complex-1-signaling-is-required-for-the-steady-state-in-vivo-development-of-inflammatory-monocytes/