Production of Extracellular Traps against and in Infected Lung Tissue Is Dependent on Invading Neutrophils and Influenced by Hydrophobin RodA

English version České info

Aspergillus fumigatus is the most important airborne fungal pathogen causing life-threatening infections in immunocompromised patients. Macrophages and neutrophils are known to kill conidia, whereas hyphae are killed mainly by neutrophils. Since hyphae are too large to be engulfed, neutrophils possess an array of extracellular killing mechanisms including the formation of neutrophil extracellular traps (NETs) consisting of nuclear DNA decorated with fungicidal proteins. However, until now NET formation in response to A. fumigatus has only been demonstrated in vitro, the importance of neutrophils for their production in vivo is unclear and the molecular mechanisms of the fungus to defend against NET formation are unknown. Here, we show that human neutrophils produce NETs in vitro when encountering A. fumigatus. In time-lapse movies NET production was a highly dynamic process which, however, was only exhibited by a sub-population of cells. NETosis was maximal against hyphae, but reduced against resting and swollen conidia. In a newly developed mouse model we could then demonstrate the existence and measure the kinetics of NET formation in vivo by 2-photon microscopy of Aspergillus-infected lungs. We also observed the enormous dynamics of neutrophils within the lung and their ability to interact with and phagocytose fungal elements in situ. Furthermore, systemic neutrophil depletion in mice almost completely inhibited NET formation in lungs, thus directly linking the immigration of neutrophils with NET formation in vivo. By using fungal mutants and purified proteins we demonstrate that hydrophobin RodA, a surface protein making conidia immunologically inert, led to reduced NET formation of neutrophils encountering Aspergillus fungal elements. NET-dependent killing of Aspergillus-hyphae could be demonstrated at later time-points, but was only moderate. Thus, these data establish that NET formation occurs in vivo during host defence against A. fumigatus, but suggest that it does not play a major role in killing this fungus. Instead, NETs may have a fungistatic effect and may prevent further spreading.

Vyšlo v časopise: Production of Extracellular Traps against and in Infected Lung Tissue Is Dependent on Invading Neutrophils and Influenced by Hydrophobin RodA. PLoS Pathog 6(4): e32767. doi:10.1371/journal.ppat.1000873
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1000873

Souhrn

Zdroje

1. BalloyV

ChignardM

2009 The innate immune response to Aspergillus fumigatus. Microbes Infect 11 919 927

2. BrakhageAA

2005 Systemic fungal infections caused by Aspergillus species: epidemiology, infection process and virulence determinants. Curr Drug Targets 6 875 886

3. TekaiaF

LatgeJP

2005 Aspergillus fumigatus: saprophyte or pathogen? Curr Opin Microbiol 8 385 392

4. BehnsenJ

NarangP

HasenbergM

GunzerF

BilitewskiU

2007 Environmental Dimensionality Controls the Interaction of Phagocytes with the Pathogenic Fungi Aspergillus fumigatus and Candida albicans. PLoS Pathog 3 e13 doi:10.1371/journal.ppat.0030013

5. SturtevantJ

LatgeJP

1992 Participation of complement in the phagocytosis of the conidia of Aspergillus fumigatus by human polymorphonuclear cells. J Infect Dis 166 580 586

6. BonnettCR

CornishEJ

HarmsenAG

BurrittJB

2006 Early neutrophil recruitment and aggregation in the murine lung inhibit germination of Aspergillus fumigatus conidia. Infect Immun 74 6528 6539

7. MircescuMM

LipumaL

VanRN

PamerEG

HohlTM

2009 Essential role for neutrophils but not alveolar macrophages at early time points following Aspergillus fumigatus infection. J Infect Dis 200 647 656

8. ZaremberKA

SuguiJA

ChangYC

Kwon-ChungKJ

GallinJI

2007 Human polymorphonuclear leukocytes inhibit Aspergillus fumigatus conidial growth by lactoferrin-mediated iron depletion. J Immunol 178 6367 6373

9. LessingF

KniemeyerO

WozniokI

LoefflerJ

KurzaiO

2007 The Aspergillus fumigatus Transcriptional Regulator AfYap1 Represents the Major Regulator for Defence against Reactive Oxygen Intermediates but Is Dispensable for Pathogenicity in an Intranasal Mouse Infection Model. Eukaryot Cell 6 2290 2302

10. ReevesEP

LuH

JacobsHL

MessinaCG

BolsoverS

2002 Killing activity of neutrophils is mediated through activation of proteases by K+ flux. Nature 416 291 297

11. SegalAW

2005 How neutrophils kill microbes. Annu Rev Immunol 23 197 223

12. ParisS

WysongD

DebeaupuisJP

ShibuyaK

PhilippeB

2003 Catalases of Aspergillus fumigatus. Infect Immun 71 3551 3562

13. TkalcevicJ

NovelliM

PhylactidesM

IredaleJP

SegalAW

2000 Impaired immunity and enhanced resistance to endotoxin in the absence of neutrophil elastase and cathepsin G. Immunity 12 201 210

14. BrinkmannV

ReichardU

GoosmannC

FaulerB

UhlemannY

2004 Neutrophil Extracellular Traps Kill Bacteria. Science 303 1532 1535

15. FuchsTA

AbedU

GoosmannC

HurwitzR

SchulzeI

2007 Novel cell death program leads to neutrophil extracellular traps. J Cell Biol 176 231 241

16. BrinkmannV

ZychlinskyA

2007 Beneficial suicide: why neutrophils die to make NETs. Nat Rev Microbiol 5 577 582

17. JaillonS

PeriG

DelnesteY

FremauxI

DoniA

2007 The humoral pattern recognition receptor PTX3 is stored in neutrophil granules and localizes in extracellular traps. J Exp Med 204 793 804

18. UrbanCF

ReichardU

BrinkmannV

ZychlinskyA

2006 Neutrophil extracellular traps capture and kill Candida albicans yeast and hyphal forms. Cell Microbiol 8 668 676

19. BianchiM

HakkimRA

BrinkmannV

SilerU

SegerRA

2009 Restoration of NET formation by gene therapy in CGD controls aspergillosis. Blood 114 2619 2622

20. ClarkSR

MaAC

TavenerSA

McDonaldB

GoodarziZ

2007 Platelet TLR4 activates neutrophil extracellular traps to ensnare bacteria in septic blood. Nat Med 13 463 469

21. UrbanCF

ErmertD

SchmidM

bu-AbedU

GoosmannC

2009 Neutrophil extracellular traps contain calprotectin, a cytosolic protein complex involved in host defence against Candida albicans. PLoS Pathog 5 e1000639 doi:10.1371/journal.ppat.1000639

22. FaustN

VarasF

KellyLM

HeckS

GrafT

2000 Insertion of enhanced green fluorescent protein into the lysozyme gene creates mice with green fluorescent granulocytes and macrophages. Blood 96 719 726

23. PetersNC

EgenJG

SecundinoN

DebrabantA

KimblinN

2008 In vivo imaging reveals an essential role for neutrophils in leishmaniasis transmitted by sand flies. Science 321 970 974

24. ChtanovaT

SchaefferM

HanSJ

van DoorenGG

NollmannM

2008 Dynamics of neutrophil migration in lymph nodes during infection. Immunity 29 487 496

25. StegemannS

DahlbergS

KrögerA

GerekeM

BruderD

2009 Increased susceptibility for superinfection with Streptococcus pneumoniae during influenza virus infection is not caused by TLR7-mediated lymphopenia. PLoS ONE 4 e4840 doi:10.1371/journal.pone.0004840

26. LangfelderK

JahnB

GehringerH

SchmidtA

WannerG

1998 Identification of a polyketide synthase gene (pksP) of Aspergillus fumigatus involved in conidial pigment biosynthesis and virulence. Med Microbiol Immunol (Berl) 187 79 89

27. TsaiHF

ChangYC

WashburnRG

WheelerMH

Kwon-ChungKJ

1998 The developmentally regulated alb1 gene of Aspergillus fumigatus: its role in modulation of conidial morphology and virulence. J Bacteriol 180 3031 3038

28. AimaniandaV

BayryJ

BozzaS

KniemeyerO

PerruccioK

2009 Surface hydrophobin prevents immune recognition of airborne fungal spores. Nature 460 1117 1121

29. DagueE

AlsteensD

LatgeJP

DufreneYF

2008 High-resolution cell surface dynamics of germinating Aspergillus fumigatus conidia. Biophys J 94 656 660

30. ThauN

MonodM

CrestaniB

RollandC

TronchinG

1994 Rodletless mutants of Aspergillus fumigatus. Infect Immun 62 4380 4388

31. UrbanCF

LouridoS

ZychlinskyA

2006 How do microbes evade neutrophil killing? Cell Microbiol 8 1687 1696

32. GuimaraesLH

MachadoPR

LagoEL

MorganDJ

SchrieferA

2009 Atypical manifestations of tegumentary leishmaniasis in a transmission area of Leishmania braziliensis in the state of Bahia, Brazil. Trans R Soc Trop Med Hyg 103 712 715

33. MohanC

AdamsS

StanikV

DattaSK

1993 Nucleosome: a major immunogen for pathogenic autoantibody-inducing T cells of lupus. J Exp Med 177 1367 1381

34. AsensiV

KimenoK

KawamuraI

SakumotoM

NomotoK

1989 Treatment of autoimmune MRL/lpr mice with anti-B220 monoclonal antibody reduces the level of anti-DNA antibodies and lymphadenopathies. Immunology 68 204 208

35. IshiiKJ

AkiraS

2006 Innate immune recognition of, and regulation by, DNA. Trends Immunol 27 525 532

36. BurckstummerT

BaumannC

BlumlS

DixitE

DurnbergerG

2009 An orthogonal proteomic-genomic screen identifies AIM2 as a cytoplasmic DNA sensor for the inflammasome. Nat Immunol 10 266 272

37. ChiuYH

MacmillanJB

ChenZJ

2009 RNA polymerase III detects cytosolic DNA and induces type I interferons through the RIG-I pathway. Cell 138 576 591

38. TakaokaA

WangZ

ChoiMK

YanaiH

NegishiH

2007 DAI (DLM-1/ZBP1) is a cytosolic DNA sensor and an activator of innate immune response. Nature 448 501 506

39. KimJV

KangSS

DustinML

McGavernDB

2009 Myelomonocytic cell recruitment causes fatal CNS vascular injury during acute viral meningitis. Nature 457 191 195

40. JahnB

KochA

SchmidtA

WannerG

GehringerH

1997 Isolation and characterization of a pigmentless-conidium mutant of Aspergillus fumigatus with altered conidial surface and reduced virulence. Infect Immun 65 5110 5117

41. LevitzSM

SelstedME

GanzT

LehrerRI

DiamondRD

1986 In vitro killing of spores and hyphae of Aspergillus fumigatus and Rhizopus oryzae by rabbit neutrophil cationic peptides and bronchoalveolar macrophages. J Infect Dis 154 483 489

42. MonodM

ParisS

SanglardD

Jaton-OgayK

BilleJ

1993 Isolation and characterization of a secreted metalloprotease of Aspergillus fumigatus. Infect Immun 61 4099 4104

43. WozniokI

HornbachA

SchmittC

FroschM

EinseleH

2008 Induction of ERK-kinase signalling triggers morphotype-specific killing of Candida albicans filaments by human neutrophils. Cell Microbiol 10 807 820