Subversion of neutrophil apoptosis by Staphylococcus aureus: consequences for bacterial pathogenesis and host defence
Lead Research Organisation:
University of Sheffield
Department Name: Respiratory Medicine
Abstract
Staphylococcus aureus is a bacterium causing a range of serious infections, which are increasingly difficult to treat because of resistance of many strains of this bacterium to antibiotics (MRSA strains). An alternative approach to treatment is to reinforce the body‘s own defences and increase our ability to kill the bacteria. Our most important defences against Staphylococcus aureus are white blood cells called neutrophils. Neutrophils eat bacteria and then kill them using a battery of toxic enzymes. This killing of bacteria is typically followed by suicide of the neutrophils, which acts as a safe disposal mechanism for both the dead bugs and the neutrophil‘s powerful enzymes.
We found Staphylococcus aureus can prevent this neutrophil suicide process called apoptosis. Instead the neutrophils disintegrate before having killed the bacteria, releasing live bacteria so the infection continues. The purpose of this research is to determine how Staphylococcus prevents neutrophils from completing their suicide programme and to identify ways of preventing neutrophil disintegration or necrosis. Preventing the necrotic death of infected neutrophils could prolong their lifespan and increase their effectiveness at killing bacteria. We aim to identify inhibitors of the death process and thus a novel avenue of treatment for this infection.
We found Staphylococcus aureus can prevent this neutrophil suicide process called apoptosis. Instead the neutrophils disintegrate before having killed the bacteria, releasing live bacteria so the infection continues. The purpose of this research is to determine how Staphylococcus prevents neutrophils from completing their suicide programme and to identify ways of preventing neutrophil disintegration or necrosis. Preventing the necrotic death of infected neutrophils could prolong their lifespan and increase their effectiveness at killing bacteria. We aim to identify inhibitors of the death process and thus a novel avenue of treatment for this infection.
Technical Summary
Staphylococcus aureus is a major human pathogen that is increasingly difficult to treat because of rising antibiotic resistance. An alternative therapeutic strategy is to augment our host defences against the organism, with neutrophils being particularly important. We have demonstrated that S. aureus can subvert neutrophilic host defences. Following ingestion by neutrophils, S. aureus can first delay neutrophil apoptosis, allowing intracellular survival of bacteria, then induce neutrophil necrosis, resulting in release of viable bacteria. I hypothesise that strategies to re-engage the programme of apoptosis in neutrophils will prevent bacterial survival and dissemination and thus improve the outcome of S. aureus infection. My aims are, therefore, to:
1. Establish the kinetics of altered neutrophil death in S. aureus infection, quantifying apoptosis by flow cytometry, using a range of specific markers, and infection by both flow cytometry (using a GFP-S. aureus) and colony counting methods.
2. Identify S. aureus components that delay neutrophil apoptosis, by study of both candidate molecules, e.g. peptidoglycan and lipoteichoic acid, and a range of S. aureus mutants available in the Foster lab. I shall investigate the downstream signalling pathways that suppress engagement of the apoptosis programme by western blotting and use of pharmacological inhibitors.
3. Determine the mechanisms and consequences of S. aureus induced neutrophil necrosis. I shall study the effects of inhibiting known pathways of neutrophil necrosis, e.g. generation of reactive oxygen intermediates and release of lysosomal proteases. Where compounds are identified that prevent neutrophil necrosis and induce apoptosis I shall test whether these manipulations also reduce bacterial survival.
4. Test the effects of bacterial and host factors modifying neutrophil death on in vivo outcomes of S. aureus infection in a murine model of cutaneous abscess formation. These experiments will allow me to test the effects of bacterial strains deficient in regulators of neutrophil death and of compounds driving neutrophil apoptosis, evaluating the extent of neutrophilic inflammation and effects upon bacterial survival.
Enhancement of the anti-Staphylococcal actions of neutrophils by manipulation of the timing and mode of neutrophil death could result in substantial gain of treatment of this antibiotic resistant infection.
1. Establish the kinetics of altered neutrophil death in S. aureus infection, quantifying apoptosis by flow cytometry, using a range of specific markers, and infection by both flow cytometry (using a GFP-S. aureus) and colony counting methods.
2. Identify S. aureus components that delay neutrophil apoptosis, by study of both candidate molecules, e.g. peptidoglycan and lipoteichoic acid, and a range of S. aureus mutants available in the Foster lab. I shall investigate the downstream signalling pathways that suppress engagement of the apoptosis programme by western blotting and use of pharmacological inhibitors.
3. Determine the mechanisms and consequences of S. aureus induced neutrophil necrosis. I shall study the effects of inhibiting known pathways of neutrophil necrosis, e.g. generation of reactive oxygen intermediates and release of lysosomal proteases. Where compounds are identified that prevent neutrophil necrosis and induce apoptosis I shall test whether these manipulations also reduce bacterial survival.
4. Test the effects of bacterial and host factors modifying neutrophil death on in vivo outcomes of S. aureus infection in a murine model of cutaneous abscess formation. These experiments will allow me to test the effects of bacterial strains deficient in regulators of neutrophil death and of compounds driving neutrophil apoptosis, evaluating the extent of neutrophilic inflammation and effects upon bacterial survival.
Enhancement of the anti-Staphylococcal actions of neutrophils by manipulation of the timing and mode of neutrophil death could result in substantial gain of treatment of this antibiotic resistant infection.
