Transcript Metabolismus leukocytů a trombocytů

Metabolism of leukocytes
and platelets
[email protected]
Differentiation of the bone marrow stem cells
stem cell
myeloid
progenitor
megakaryocyte
erythroblast
lymphoid
progenitor
erythrocyte
monocyte
platelets
neutrophil eosinophil basophil
dendritic cell
macrophage
mature
lymphocytes
plasma
cell
A) Phagocytic cells:




Neutrophils – most abundant
Eosinophils
Monocytes
Macrophages – rise by differentiation of monocytes in tissues
Degradation of the ingested particle:
 1) Activation of NADPH oxidase
 2) Production of NO by nitric oxide synthase
 3) Fusion of phagosome with lysosomes of the phagocytic cell that
contain bactericidal substances and hydrolytic enzymes (often with
acidic pHopt)
1) NADPH-oxidase
 Protein complex of neutrophils, eosinophils, monocytes, macrophages
 NADPH + 2 O2 → NADP+ + H+ + 2 O2•- superoxide anion
2 O2•- + 2 H+ → O2 + H2O2
 H2O2 can damage bacteria directly or after conversion to OH• :
H2O2 + M+ → OH• + OH- + M2+ (M; metal)
 Activation: by association of the components localized in cytosol with
cytochrome b558 in the membrane; electrons from cytosolic NADPH
are – via FAD and cytochrome – transferred to oxygen
cytochrome b558
active NADPH-oxidase
plasma
membrane
fusion
with
lysosomes
phagosome
Myeloperoxidase
 Present in granules of neutrophils and monocytes, but not macrophages!
 Significant portion of H2O2 (produced by dismutation of O2•- generated by
NADPH oxidase) is used by myeloperoxidase to oxidize Cl- to HClO
 HClO is highly reactive, able to oxidize biomolecules; it also provides
toxic chlorine gas:
HClO + H+ + Cl- → Cl2 + H2O
 HClO also reacts with O2•- yielding OH•:
HClO + O2•- → O2 + OH• + Cl-
Chronic granulomatous disease
 Caused by a deficiency of one of the NADPH oxidase subunits
 Superoxide and the other reactive oxygen species are not produced
 Severe infections that are very hard to treat – e.g.:
 Burkholdaria cepacea causes pneumonia
 Aspergillus causes intractable pneumonia, septicaemia; can lead
to death
 Treatment: antibiotics, antifungal agents
2) Nitric oxide production
 Mainly by inducible nitric oxide synthase (iNOS) of macrophages which is
induced by cytokines (INF-γ, TNF) or bacterial lipopolysaccharide:
Arg
citrulline
 NO• can kill bacteria directly (e.g. by inhibition of the respiratory chain) or
indirectly: by reaction with O2•-, generating peroxynitrite ONOO- which
attacks Fe-S proteins and essential –SH groups, inactivates enzymes…
 NADPH oxidase is effective mainly in degradation of extracellular
pathogens (Salmonella, Staphylococcus, Streptococcus
pyogenes)…neutrophils
X
 NO serves mainly to kill the intracellular parasites (Listeria, Brucella,
Candida albicans)…macrophages
3) Granules (lysosomes) of neutrophils
 Contain bactericidal substances and hydrolases that, after fusion with
phagosome, destroy the engulfed particles:
 myeloperoxidase
 lysozyme – cleaves glycosidic bonds in peptidoglycan of the bacterial
(primarily G+) cell walls
 defensins – cationic peptides (Arg) with Mr of 3,5-6 kDa; interact with
anionic lipids of bacterial membrane and make pores in it; can also
inhibit synthesis of DNA and proteins
 hydrolases, e.g. elastase – serine protease: can damage bacteria
and cleave virulence factors, but also cause harm to host tissues
(cleaves the proteins of extracellular matrix, too)
Eosinophils
 Main task: defence against multicellular parasites
 Display all the above-mentioned mechanisms with slight differences:
 ROS production
 peroxidase of eosinophils – similar to myeloperoxidase, but prefers
Br- as a substrate (instead of Cl-), thus generating HBrO (instead of
HClO)
 basic protein of eosinophils disrupting the parasite cell membranes
B) Basophils and mast cells
 Activated by antigens / allergens interacting with IgE bound to the
surface IgE receptors of basophils (mast cells)
 Upon activation, content of their granules is released – substances that
are harmful to parasite and induce reactions that should lead to its
removal; however, they can also be responsible for allergic symptoms:
 hydrolases
 histamine
 heparin
cytoplasmic granules
 Synthesis of eicosanoids is activated; leukotrienes are potent
bronchoconstrictors, stimulate chemotaxis and leukocyte activation
Histamine
 Produced by histidine decarboxylation:
 Causes vasodilation and bronchoconstriction  helps to eliminate
parasites (cough, peristalsis, enhanced production of mucus)
Atopy
 IgE recognizing allergens (from pollen, food…) are produced and bind to
IgE receptors of basophils (mast cells). Next exposure to the allergen can
lead to release of histamine and heparin and synthesis of eicosanoids
 Local symptoms occur: allergic rhinitis, asthma, conjunctivitis
 If the allergen enters bloodstream, it can cause a massive degranulation
of basophils (mast cells)  increase in vascular permeability, decrease in
blood pressure  pulmonary oedema, ischemia… anaphylactic shock
 Treatment: antihistamines – block histamine receptors
C) Lymphocytes
 Have specific receptors recognizing one particular antigen: B cell
receptors (BCR) and T cell receptors (TCR), respectively
 BRC is a membrane-bound immunoglobulin, TCR is very similar to Ig
 B cells (after proliferation and differentiation into plasma cells) secrete
large amounts of antibodies (soluble immunoglobulins)
Soluble immunoglobulins
VH
VL
CH1
Fab
CL
CH2
Fc
CH3
 2 heavy chains (H) interconnected by
disulfide bonds
 2 light chains (L), each connected to
one of the H chains (by disulfide bond)
 H chain: 4-5 domains, 50-75 kDa
L chain: 2 domains, 25 kDa
 N-terminal domains of H- and Lchains are variable (VH resp. VL), the
others are constant (CH resp. CL), i.e.
the same in one type of Ig
 Variable domains of H a L chains form
the antigen-binding site
Types of immunoglobulins (Ig)
155 kDa
(similar: IgD, IgE)
900 kDa
 There are 2 isotypes of L: κ, λ
 There are 5 isotypes of H:
, γ, δ, ε, μ
 According to these isotypes of H,
5 types of immunoglobulins can
be distinguished:
 IgA (2 subtypes)
 IgG (4 subtypes)
 IgD
 IgE
 IgM
 IgM can form pentamer, IgA can
form dimer or trimer
D) Platelets
 No nucleus  many of their metabolites come from megakaryocytes
 Form blood clots, act as vasoconstrictors
 Participate in defence against infections, e.g.: they suppress the growth
of Plasmodium falciparum (infectious agent that causes malaria)
 Generate O2•- and H2O2 that may synergize with pro-aggregatory stimuli
 Contain thromboxan A synthase that catalyzes conversion of prostaglandin H2 to thromboxan A2:
TXA2 – promotes platelet aggregation and vasoconstriction
 Platelets also release two very important factors that can influence
not only platelets but also other cell types:
 Platelet-Activating Factor (PAF)
 Platelet-Derived Growth Factor (PDGF)
Platelet-Activating Factor
phospholipid
 Mainly juxtacrine and paracrine signalling via GPCR
 Promotes platelet aggregation
 Induces activation of leukocytes, adhesion, chemotaxis, cytokine
production, causes vasodilation and bronchoconstriction
 Mediates interplay between thrombotic and inflammatory cascades
 BUT: it is also suspected of contributing to allergy, anaphylactic shock…
 It is produced also by endothelial cells, monocytes, granulocytes…
Platelet-Derived Growth Factor
 Dimeric protein, 3 isoforms
 Receptors: tyrosine kinases – expressed on fibroblasts, glia, smooth
muscle cells, leukocytes….
 Effects:
 proliferation
 chemotaxis
 cytoskeletal rearrangements
 differentiation of certain types of cells (e.g. in CNS)
  participates in wound healing, capillary formation, embryonic and
postnatal development!
 BUT: probably also plays a role in pathogenesis (some tumours)
Cytokines
 Proteins secreted by leukocytes and other cells (but there are also
membrane cytokines) that influence (via receptors) the cells of the
immune system
 Cytokine signalling:
 autocrine – a cytokine influences the same cell that produces it
 paracrine – a cytokine influences the nearby cells
 endocrine – a cytokine influences distant cells (after transport by the
bloodstream)
Types of cytokines
 Interleukins – e.g. IL-6: produced by macrophages, neutrophils, stimulates lymphocytes, secretion of Ig, synthesis of acute phase reactants
 Chemokines – induce chemotaxis
 Interferons – e.g. INF-: produced by lymphocytes, monocytes, and
macrophages, participates in antiviral defense (induces synthesis of
enzymes that block viral replication)
 Transforming growth factors – e.g. TGF-β: produced by T-lymphocytes,
macrophages, and platelets, displays anti-inflammatory effects
 Tumor necrosis factors – e.g. TNF-β: able to induce apoptosis
Leukocyte infiltration into tissues
= diapedesis (extravasation):
Taken from:
Halliwell, Gutteridge,
Oxford University Press, 1999
 Leukocytes are slowed down by the interaction of their mucins with selectines on
the surface of endothelial cells (EC)
 Cytokines on the surface of EC interact with the receptors of leukocytes
 A strong adhesion mediated by the interaction of integrins with molecules on the
surface of EC → migration of leukocytes into the tissue directed by cytokins
released by inflammatory cells or EC
Regulation
 Many functions of leukocytes are regulated by monomeric GTP-binding
proteins, e.g. Rac, Rho:
 activation of NADHP oxidase
 chemotaxis
 phagocytosis
 fusion of phagosome with granules
 Rho and Rac are able to modulate the assembly of actin filaments,
which plays a role in the processes listed above

http://uk.video.search.yahoo.com/video/play?ei=UTF-8&fr=yfp-t702&p=chemotaxis&vid=0001539076618&dt=&l=77&turl=http%3A%2F%2Fyts.video.search.yahoo.com%2Fimage%2F2021a80a1&rurl=http%3A%2F%2Fwww.youtube.com%2Fv
%2FZUUfdP87Ssg%26hl%3Den%26fs%3D1&tit=Neutrophil