Toll-like receptor 2 ligands on the staphylococcal cell wall downregulate superantigen-induced T cell activation and prevent toxic shock syndrome Thu A Chau 1 , Michelle L McCully 1,2 , William Brintnell 3 , Gary An 4 , Katherine J Kasper 2 , Enrique D Vine ´s 2 , Paul Kubes 5 , S M Mansour Haeryfar 2 , John K McCormick 2,6 , Ewa Cairns 2,3 , David E Heinrichs 2 & Joaquı ´n Madrenas 1–3 Staphylococcal superantigens are pyrogenic exotoxins that cause massive T cell activation leading to toxic shock syndrome and death. Despite the strong adaptive immune response induced by these toxins, infections by superantigen-producing staphylococci are very common clinical events. We hypothesized that this may be partly a result of staphylococcal strains having developed strategies that downregulate the T cell response to these toxins. Here we show that the human interleukin-2 response to staphylococcal superantigens is inhibited by the simultaneous presence of bacteria. Such a downregulatory effect is the result of peptidoglycan-embedded molecules binding to Toll-like receptor 2 and inducing interleukin-10 production and apoptosis of antigen-presenting cells. We corroborated these findings in vivo by showing substantial prevention of mortality after simultaneous administration of staphylococcal enterotoxin B with either heat-killed staphylococci or Staphylococcus aureus peptidoglycan in mouse models of superantigen-induced toxic shock syndrome. S. aureus is a common human commensal organism, present in the upper respiratory tract of 25–50% of the population 1 . It is also one of the most frequent pathogens, being linked to almost 500,000 hospi- talizations per year in the United States alone 2,3 . The high morbidity associated with S. aureus has further increased with the appearance of strains with broad antibiotic resistance, including methicillin-resistant strains responsible for up to 20,000 deaths per year 3 . These facts make S. aureus a considerable healthcare problem, with an estimated cost at $9 billion per year in the United States 3,4 . S. aureus is a major source of superantigens 5 . These toxins can activate up to 20% of all T cells in the body by binding the human leukocyte antigen (HLA) class II molecules on antigen-presenting cells (APCs) and specific V b regions of the T cell receptor (TCR) 6,7 . Between 50% and 80% of S. aureus isolates are positive for at least one superantigen gene 8–11 , and close to 50% of these isolates show superantigen production and toxin activity 12 . Given the high frequency of infections by superantigen-producing S. aureus and the potent immunostimulatory capacity of superanti- gens, one might expect that staphylococcal toxic shock syndrome (TSS) would be a rather common event. However, epidemiological studies do not support this idea. It was initially estimated that the incidence of menstrual TSS by S. aureus is around 6.2 cases per 100,000 women 12 to 49 years of age 13 . This figure has been steadily decreasing to the current number of 1 case per 100,000 women aged 18 to 44 years 14 . The incidence of other types of staphylococcal TSS shows a similar pattern and varies from 0.3 to 3.2 cases per 100,000, depending on age and clinical setting. The relatively low frequency of staphylococcal TSS despite the high frequency of infections by superantigen-producing S. aureus remains unexplained. It is probably not due to lack of exposure to super- antigens, because humoral immunity against these exotoxins is detected in almost 100% of the population by the fifth decade of life 11,15 . One reason may be that superantigens are not produced as much in blood as in tissues, owing to inhibitory properties of the a- and b-globin chains 16 . But we have noticed that bacteremia is rarely detected in individuals with staphylococcal TSS, in contrast to those with staphylococcal sepsis who have positive blood cultures but no TSS 17–19 . Indeed, a positive blood culture is not required to diagnose staphylococcal TSS 20 . Given that the presence of superantigens in the body would often be concomitant with the presence of the staphylo- cocci producing them, we reasoned that the relatively low frequency of staphylococcal superantigen-induced TSS might be partly due to downregulation of the T cell response by the bacteria. Here we show that the cell wall of S. aureus and other Gram- positive bacteria contains peptidoglycan-embedded molecules that act as Toll-like receptor 2 (TLR2) ligands and inhibit the interleukin-2 Received 12 November 2008; accepted 7 April 2009; published online 24 May 2009; doi:10.1038/nm.1965 1 The FOCIS Centre for Clinical Immunology and Immunotherapeutics, Robarts Research Institute, London, Ontario, Canada. 2 Department of Microbiology and Immunology, The University of Western Ontario, London, Ontario, Canada. 3 Department of Medicine, The University of Western Ontario, London, Ontario, Canada. 4 Department of Surgery, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA. 5 Department of Physiology and Biophysics, University of Calgary, Calgary, Alberta, Canada. 6 Lawson Health Research Institute, London, Ontario, Canada. Correspondence should be addressed to J.M. (madrenas@robarts.ca). NATURE MEDICINE VOLUME 15 [ NUMBER 6 [ JUNE 2009 641 ARTICLES © 2009 Nature America, Inc. All rights reserved.