درمان مننژیت به کمک آنزیم DNase
مننژیت یک بیماری مهلک با نرخ مرگ و میر بالا است، برای مثال این بیماری عامل اصلی مرگ کودکان در هند است. Streptococcus pneumonia عامل نوع باکتریایی این بیماری است.
مواجه سیستم عصبی انسان با این باکتری موجب فعال شدن نوتروفیلها و به دنبال آن تشکیل شبکههای خارج سلولی (NETs) میشود که باکتریها را به دام میاندازد. تجمع این تودههای باکتریایی در مایع مغزی نخاعی بیماران یکی از عوامل مصنونیت آنها در برابر درمان آنتی بیوتیکی است. یافتههای جدید نشان میدهد استفاده همزمان از آنزیم DNase I در کنار آنتی بیوتیک موجب افزایش کارایی درمان میشود. DNase موجب پاره شدن شبکههای خارج سلولی و به دنبال آن دسترسی آنتیبیوتیک به باکتریها خواهد شد.
Neutrophil extracellular traps in the central nervous system hinder bacterial clearance during pneumococcal meningitis
Neutrophils are crucial mediators of host defense that are recruited to the central nervous system (CNS) in large numbers during acute bacterial meningitis caused by Streptococcus pneumoniae. Neutrophils release neutrophil extracellular traps (NETs) during infections to trap and kill bacteria. Intact NETs are fibrous structures composed of decondensed DNA and neutrophil-derived antimicrobial proteins. Here we show NETs in the cerebrospinal fluid (CSF) of patients with pneumococcal meningitis, and their absence in other forms of meningitis with neutrophil influx into the CSF caused by viruses, Borrelia and subarachnoid hemorrhage. In a rat model of meningitis, a clinical strain of pneumococci induced NET formation in the CSF. Disrupting NETs using DNase I significantly reduces bacterial load, demonstrating that NETs contribute to pneumococcal meningitis pathogenesis in vivo. We conclude that NETs in the CNS reduce bacterial clearance and degrading NETs using DNase I may have significant therapeutic implications.
Acute bacterial meningitis (ABM) caused by Streptococcus pneumoniae(pneumococci) is a life-threatening medical condition that is accompanied by a high risk of debilitating neurological sequelae in survivors1,2,3. Pneumococci are not only one of the most frequent but are also one of the most lethal pathogens to cause ABM in both adults and children. Despite the introduction of antibiotics and vaccines, the mortality rates associated with pneumococcal ABM remains high4,5. The emergence of antibiotic-resistant strains and nonvaccine serotypes of pneumococci have further complicated therapy6.
The onset of bacterial infection results in the activation of both noncellular and cellular components of the immune system, with neutrophils among the first immune cells that are actively recruited to sites of infections7. The central nervous system (CNS) lacks extensive neutrophil immune-surveillance during healthy conditions, while during ABM a massive neutrophil recruitment occurs across the blood brain barrier (BBB) to eliminate bacteria8. In fact, the usually clear cerebrospinal fluid (CSF) takes on a turbid cloudy appearance due to the overwhelming presence of neutrophils and bacteria during pneumococcal ABM9.
The primary function of the neutrophils is to rapidly engage and clear invading pathogens. Neutrophils execute their function by phagocytosis, producing reactive oxygen species (ROS), hypochlorite and releasing antimicrobial protein/peptides through degranulation7. In addition, neutrophils have been described to resist invading bacteria by secreting decondensed nuclear DNA coated with granule-derived antimicrobial proteins called neutrophil extracellular traps (NETs) into the external environment10. NET formation is an evolutionarily conserved innate immune response that is directed at capturing and killing microbial pathogens11. The neutrophil-derived cationic proteins and proteases that coat NETs exhibit potent antimicrobial activity against a wide variety of microbial pathogens12. Recently, it was also reported that short DNA fragments derived from the DNA-backbone of NETs can also exert bactericidal effects against Pseudomonas aeruginosa and Staphylococcus aureus13. The role of NETs as an antibacterial strategy is only beginning to unravel and may be dependent on the type and site of infection14,15,16,17.
In spite of their antimicrobial properties, the excessive presence of NETs can be detrimental to the host in some cases, and hence NETs have been aptly acknowledged as “double-edged swords of innate immunity”۱۸. NET-components such as extracellular DNA and antimicrobial proteins, including histones and neutrophil elastase (NE)10, are proinflammatory and contribute to disease severity19,20. Since NETs were discovered to have antimicrobial effects over a decade ago, several reports have demonstrated the prevalence of NET evasion mechanisms in bacteria, including pneumococci21. The pneumococci can evade NET-mediated killing by degrading NET-DNA secreting nucleases22,23, or by incorporating positively charged D-alanine into surface lipoteichoic acids, enabling the bacteria to repel cationic antimicrobial peptides bound to NETs24. These mechanisms allow pneumococci to remain trapped within NETs without being killed and aid in their dissemination from the lungs into the blood stream21.
To date, the presence of NETs has been demonstrated in the CSF in a single study by using a porcine model of Streptococcus suis meningitis25. However, the presence of NETs in human CSF has never been established. We undertook the study to investigate whether NETs are formed in CSF during pneumococcal infections and, if present, to characterize their functional importance in ABM. Here, we demonstrate the presence of NETs and NET-related proteins in the CSF of pneumococcal ABM patients using immunofluorescence microscopy and mass spectrometry. A rat model of pneumococcal meningitis further revealed neutrophil recruitment and NET formation in the CSF. The presence of NETs in the CSF compartment was found to promote pneumococcal survival and dissemination into other organs. Degradation of NET-associated DNA using DNase I resulted in the clearance of bacteria from the brain, lungs, spleen and blood of infected animals. This was because DNase treatment increased oxidative burst and phagocytosis in neutrophils coincubated with pneumococci. Our data indicate that NET formation in the CSF during pneumococcal meningitis might be detrimental to the host and that degrading these structures using DNase I leads to enhanced bacterial clearance from the brain.