<i>Staphylococcus aureus</i> toxins mediate endothelial Thrombomodulin release during severe invasive infections

Thrombomodulin (TM) is a membrane protein with significant roles in coagulation hemostasis and immune response. Its soluble form (sTM) has recently emerged as a key biomarker for severe invasive bacterial infections, including Necrotizing Soft Tissue Infections (NSTI). While various mechanical, chemical, and enzymatic mechanisms have been linked to TM shedding, this study investigates the direct impact of bacterial stimuli on soft tissue cells as primary sources of TM release. We stimulated organotypic models, composed of fibroblast and endothelial cells, with NSTI clinical isolates and found that while Group A Streptococcus and <i>Escherichia coli</i> had minimal effect on TM release, <i>Staphylococcus aureus</i> infection triggered a significant increase of sTM levels. We further assessed whether the secreted proteins of <i>S. aureus</i> led to higher TM levels by increased expression, increased cell toxicity, or direct cleavage of TM from the endothelial cell membrane. To investigate these mechanisms, we performed <i>in vitro</i> stimulations of endothelial monolayers with secreted proteins of two <i>S. aureus</i> isolates differing in their <i>agr</i>-system functionality. Our results indicate that <i>S. aureus agr</i>-regulated proteins induce TM shedding by direct cleavage from the cell membrane, an effect that was inhibited by metalloproteinase inhibitors. Stimulation with the pore-forming protein α-toxin showed similar results, suggesting a potential involvement of ADAM10 in TM cleavage. Additionally, we observed that other <i>agr</i>-regulated proteins can cleave TM directly. Altogether, this study reveals a pathogen-specific mechanism for TM release during <i>S. aureus</i> invasive infection, contributing to its elevated plasma levels and providing deeper insights into the pathophysiology of NSTI.