TLR2 or TLR4 Stimulation Induces Transmembrane TNF-Driven Priming in Macrophages Which Results in Improved Clearance of a Subsequent Staphylococcus aureus Infection

Abstract

Introduction: Toll-like receptor (TLR) engagement on macrophages can improve responsiveness to infection. TNF is upregulated following TLR2 or TLR4 stimulation. We sought to determine whether and how the two bioactive forms of TNF, soluble (sTNF) and transmembrane (tmTNF), may be contributing to macrophage priming, which improved responsiveness to subsequent Staphylococcus aureus infection.

Methods: RNA sequencing and cytokine quantification assays identified differentially upregulated cytokines in response to TLR2 stimulation. Immortalized and primary bone marrow-derived macrophages (BMDMs) coupled with receptor blocking and cytokine supplementation were used to investigate whether/how prior TLR-primed macrophages improved S. aureus clearance.

Results: TLR2 or TLR4 stimulated TNF-/- BMDMs failed to efficiently clear a subsequent S. aureus infection compared to TLR-stimulated wild-type (WT) BMDMs. Depletion of sTNF from TLR-stimulated WT BMDMs retained their improved S. aureus clearance. Exogenous sTNF supplementation to TNF-/- BMDMs did not rescue improved S. aureus clearance. Cell density assays showed cell-to-cell contact was important for TLR-induced improvement of S. aureus clearance. Conversely, blocking TNFR2 reduced BMDM clearance of S. aureus, despite TLR2 stimulation.

Conclusions: Our results demonstrated that TNF produced in response to TLR stimulated BMDMs was required for improved clearance of a subsequent S. aureus infection. We found that sTNF did not contribute to this priming, which suggested that tmTNF may be critical for BMDM priming which leads to improved S. aureus clearance.

Keywords: Bacteria; Macrophage; Staphylococcus aureus; Toll-like receptor; Transmembrane TNF; Tumor necrosis factor.