Differential regulation of monocyte exhaustion by distinct TLR agonists and underlying mechanisms

Monocyte exhaustion is a dysfunctional immune state marked by persistent inflammation and immune suppression, reflected in STAT1-mediated expression of pathogenic inflammatory mediator CD38 and immune suppressive PD-L1, as well as a suppression of Akt signaling and a reduction of immune-enhancing mediator CD86. While prolonged Toll-like receptor 4 (TLR4) stimulation induces monocyte exhaustion, the roles of other TLRs remain unclear. Here, we systematically evaluated the ability of TLR-2, TLR-3, TLR-7, and TLR-9 agonists to induce murine bone marrow-derived monocytes exhaustion in vitro. Although all tested agonists promoted exhaustion phenotypes to varying degrees, characterized by upregulation of STAT1 mediated expression of CD38 and PD-L1, only TLR-2 and TLR7 agonists drastically suppressed Akt and CD86. In contrast, TLR-3 or TLR-9 agonists preferentially sustained Akt activation and CD86 expression. Mechanistically, STAT1/STAT3 activation mediated by mTROC1 was common across TLR agonists responsible for elevated expression of CD38 and PD-L1. On the other hand, Akt signaling mediated by mTORC2 responsible for the expression of CD86 was preferentially suppressed by TLR-2 and TLR-7 agonists, but retained by TLR-3 and TLR-9 agonists. Deletion of Rictor, a key component of mTORC2, blocked the activation of Akt/CD86 triggered by TLR-3/9 agonists, and further elevated mTORC1 mediated activation of STAT1/3 as well as CD38 expression. Conversely, Fumagillin or Rapamycin treatment, which has been associated with reduced mTORC1 signaling activity, mitigated TLR-2/7-induced STAT1/STAT3 activation and CD38 expression. These findings reveal that monocyte exhaustion is a shared but differentially regulated outcome of distinct TLR pathways, with the mTOR axis potentially serving as a key therapeutic target for immune dysfunction.