Neutrophils in tb

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Neutrophils in tb

  1. 1. WELCOME K.KARTHIK, MVSc, VBM, IVRI
  2. 2. 4000 A DAY!!! M.tuberculosis when unleashed, lashes 4000/day
  3. 3. INTRODUCTION • TB is transmitted through aerosol – macrophages and dendritic cells first to encounter bacilli • Participation of neutrophils in monocyte recruitment, granuloma formation and lung repair (Antony et al., 1983)
  4. 4. FACTS ABOUT NEUTROPHILS • Kills organism by both oxidative (phagocytic) & non oxidative ( degranulation ) ( Kumar et al., 2010) GRANULE TYPE PROTEIN azurophilic granules (or "primary granules") myeloperoxidase, bactericidal /permeability-increasing protein (BPI), Defensins, and the serine proteases neutrophil elastase specific granules (or "secondary granules" Lactoferrin and Cathelicidin tertiary granules cathepsin and gelatinase
  5. 5. NEGLECTED NEUTROPHILS • Poorly ranked in case of TB Short lived Easily activated Cryopresevation difficult Invitro study difficult
  6. 6. NEED OF THIS TOPIC? • Commonly affected phagocyte in human Eum et al., 2010 • Contribute to control of TB in bloodi Mirtineau et al., 2007 • Neutrophil driven interferon- disease pathogenesis Berry et al., 2010
  7. 7. NEUTROPHILS IN TB – 2 WAY TRACK TRACK 1 • Neutrophils causes decrease in downstream CFU in lung – with 200 M.tuberculosis infection ( Sugarwara et al., 2004) • Depleting murine granulocyte before challenge with infection , increase CFU ( Barrios et al., 2006) TRACK 2 • No effect on CFU in case of granuloctye depletion ( Seiler et al., 2000) • RB6-8C5 monoclonal antibody – to deplete granulocyte receptor , also targets dendritic and monocytes ( Wojtasiak et al., 2010)
  8. 8. BACK TO THE BASICS Preliminary steps by neutrophils after entry of the organism: • Recruitment • Recognition • Phagocytosis • Killing
  9. 9. NEUTROPHIL RECRUITMENT PACE OF RECRUITMENT (HOURS) PLACE ORGANISM REFERENCE 1 Multiple perivascular sites M.tuberculosis Long et al., 1931 2 Hepatic infiltration M.avium Feng et al., 2003 3 Skin infiltration BCG , rabbits Shigenaga et al., 2001 4 Dermal infiltration BCG , mice Abadie et al., 2005
  10. 10. MECHANISM OF RECRUITMENT • Sensitized animals- powerful immune response to mycobacterial challenge ( Long et al., 1931) • IL 17 & IL 23 from Th 17 – masters the orchestera ( Cruz et al., 2010) • IL8 from macrophage also joins the party (Lyons et al., 2002)
  11. 11. STEPS INVOLVED Initial signal - cytokine release Activation of endothelium, increase in adhesion molecules Influx of neutrophils, initiation of complement through chemo attractant C5a
  12. 12. PHAGOCYTOSIS • Neutrophils directly interact & internalize mycobacteria ( Wolf et al., 2007) 2 mechanism mediate interaction • Direct recognitition • Opsonisation
  13. 13. DIRECT RECOGNITION • Pattern recognition receptor mediate interaction (May et al., 1987) • TLR2 also involved • Impaired control of M.tuberculosis & M.avium in TLR2 deficient mice (Feng et al., 2003) • TLR2 – mycobacterial ligand – lipoarabinomannan / 19 Kda lipoprotein (Neufert et al., 2001) • TLR4 also involved – blocking – reduce IL8 production ( Godaly et al., 2005) • Complement receptor 3&4 also bind directly (Aleman et al., 2004)
  14. 14. OPSONISATION • Opsonisation also plays important role in regulating phagocytosis • Reduction in ficoll isolated neutrophils to phagocytose after heat inactivation of serum ( Majeed et al., 1998)
  15. 15. DOES IT KILL MYCOBACTERIA? • Controversial stuff • Theoretically – it kills & halt during early infection MECHANISM: • Human neutrophil peptides – α defensin – cationic – azurophil granules bind to anionic molecules (Fu et al., 2003) • M.avium, M.kansasii, M.smegmatis, M.tuberculosis fail to bind ( Perskrist et al., 2002)
  16. 16. Cont.. • M.tuberculosis gene lysX – similar to S.aureus gene mprF – increases lysine content – decreases negative charge- decrease suseptibility to HNP ( Maloney et al., 2009) • HNP also be taken by macrophages – ability to kill organism ( Sharma et al., 2000) • Phagocytosis of apoptotic neutrophils by macrophages – restriction of mycobacterial growth ( Tan et al., 2006)
  17. 17. TROJAN HORSE • In the absence of killing the bacteria, neutrophils traffics the infection to other organs – GRANULOCYTE TROJAN HORSE ( Eruslanow et al., 2005) • Mice treated with anti IL17 during infection shows 100 fold low organism in spleen (Redford et al., 2010)
  18. 18. NEUTROPHIL EXTRACELLULAR TRAPS (NETs) • NETs composed of nuclear chromatin / mitochondrial DNA associated with histones & granular antimicrobial proteins ( Yousefi et al., 2009) • Formed in respone to pro inflammatory stimuli ( Brinkmann et al., 2004) • It traps Mycobacteria ( Ramos et al., 2009) – unable to kill – instaed it kill Listeria – confirming anti microbial property • Hence it causes localization – basis of granuloma
  19. 19. ECTOSOMES (ECTs) • Is the vesicles from cell membrane in respone to stimuli ( Gasser et al., 2003) • Ranges from 50-200 nm – have chemo attractant & pro inflammatory property • It is cholesterol enriched , express CD35 ( CR1) (Gasser et al., 2003) • ECTs from PMN bind to endothelial & macrophages but not to red cells – play role in immune response
  20. 20. NETs & ECTs NETs ECTs
  21. 21. NEUTROPHILS & MACROPHAGES CO-OPERATION • Clearance of short lived neutrophils is carried out by macrophages • Neutrophil derived chemokines attract monocyte from blood (Mantovani et al., 2011) • Mycobacterial Lipoarabinomannan – stimulate macrophage chemo taxis (Fietta et al., 2000) • Increase in apoptosis in neutrophils after mycobacterial internalization – oxidative process (Persson et al., 2008)
  22. 22. APOPTOSIS- ANTI / PROINFLAMMATORY ANTI INFLAMMATORY • Apoptosis – anti inflammatory results in induction of TGF ß, PGE2 • Inhibits IL6 ,IL8, IL12 & TNF from macrophages ( Krysko et al., 2006) PRO INFLAMMATORY • Pro inflammatory due to expression of heat shock proteins (Perrson et al., 2008) • Activation of macrophage by neutrophil proteases
  23. 23. Cont.. • Phagocytosis of apoptotic cell- may be anti / pro inflammatory based on : Mycobacteria inside neutrophil is alive / dead • Live: pro inflammatory • Dead : anti inflammatory
  24. 24. AS SIMPLE AS THAT……
  25. 25. NEUTROPHILS ON ACQUIRED IMMUNITY • Neutrophils produce IL12, interferon gamma, macrophage inflammatory protein – attracts T lymphocte ( Seiler et al., 2003) • Produce IL10 limit acquired immunity (Dorhoi et al., 2010) • Cross present antigen to dendritic cells ( Morel et al., 2008)
  26. 26. TOTAL EFFECT OF NEUTROPHILS
  27. 27. CONCLUSION • Neutrophils are seen in the early stages of the Mycobacterial infection. • In chronic cases the same neutrophils may act in the pathology of Granuloma formation • Thus neutrophil act as a “Double edged Sword”. • Whether it kills Mycobacteria ?? • It may disseminate the organism to various organs.

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