Biologyofmastcell 140714021223 Phpapp01

Biology of Mast Cell
By
Wat Mitthamsiri, MD.
Allergy and Clinical Immunology Fellow
King Chulalongkorn Memorial Hospital
Outline
Introduction
Development
Heterogeneity
Homing mechanism
Ultrastructure and mediators
Activation mechanism
Roles in allergen sensitization
Roles in allergic diseases
Introduction
History
276 million years ago
Mast cells already present in
primitive reptiles
1863, RECKLINGHAUSEN
Granulated cells found
1878/1879 EHRLICH
Mast cells and basophils
U Blank, et al., Allergy 2013; 68: 10931101.
P Bradding, H Saito., Middletons Allergy 8th edition, 2013, 228-251.
Some milestones

U Blank, et al., Allergy 2013; 68: 10931101.
Development
Development
J Douaiher, et al., Advances in Immunology, Volume 122, 2014: 211-252
Development
J Douaiher, et al., Advances in Immunology, Volume 122, 2014: 211-252
Obligate growth factor
Langley KE, et al., Blood 81 (3): 65660.
Zhang, et al., Proc.Natl.Acad.Sci.USA, 2000, 97: 7732-7737.
P Bradding, H Saito., Middletons Allergy 8th edition, 2013: 228-251.
Stem cell factor (SCF)
Other names:
Kit ligand
Steel factor
141 residues
Molecular weight
18.5 KDa
Level in normal human blood serum
3.3 ng/mL
Obligate growth factor
Geissler EN, et al., Somat. Cell Mol. Genet. 17 (2): 20714.
Stem cell factor (SCF)
Gene locus:
Chromosome 12q22-12q24 in humans
Ligand for Kit (CD117) encoded by the
proto-oncogene c-kit
Derived from many cellular sources
Epithelial cells
Mesenchymal cells
Soluble and membrane-bound forms
Other factors

Factors that enhance mast cell growth
and survival
Nerve growth factor (NGF)
IL-3
IL-6
IL-9
IL-10
Lysophosphatidic acid (LPA)
Silencing of MS4A2 gene
TRPM7 ion channel

Other factors

Factors that inhibit mast cell growth
Granulocyte-macrophage colony-
stimulating factor (GM-CSF)
Retinoids
Transforming growth factor- (TGF-)
Other factors

Factors that can either enhance or
inhibit mast cell growth and survival
IL-4
Inhibits proliferation of immature human
peripheral bloodderived mast cells (HPBMCs)
Potentiates proliferation of more mature
HPBMCs,
IL-5 and interferon-gamma (IFN-)
Prolong HCBMC survival on SCF withdrawal
Inhibit immature HPBMC proliferation
Heterogeneity
Factors leading to
heterogeneity

Interactions with the tissue matrix and
resident cells such as fibroblasts
Progenitors are possibly committed to a
particular phenotype early in their
development
Main mast cell subsets

Other mast cell subsets

Mast cells expressing tryptase and
carboxypeptidase A, but not chymase
Found in the airway epithelium in
asthmatic airways
Mast cells containing chymase and
carboxypeptidase without tryptase
(MC
C
)
Found in the lung, nose, gut, and kidney
Unknown function
Intra-tissue heterogeneity

Marked differences in:
Size and shape
Expression of
Tryptase
Chymase
FcRI
IL-9 receptor
Histidine decarboxylase
5-lipoxygenase
Leukotriene C4 (LTC4) synthase
Renin
Vascular endothelial growth factor (VEGF)
Basic fibroblast growth factor (FGF)
Plasticity
Gurish MF, Austen KF: The diverse roles of mast cells. J Exp Med 194:F1, 2001
Distinct mast cell phenotypes
in different tissues

Homing mechanism
Chemoattractants

Chemoattractants

Chemoattractants

Ultrastructure and mediators
Ultrastructure
Figure from: http://www.pathologyoutlines.com/topic/bonemarrowmastcells.html
Cell membranes contain fingerlike
projections: microplicae
Ultrastructure

Immature mast cells
May have a multilobed nucleus
Mature mast cells
Monolobed nucleus
No apparent nucleoli
Little condensed chromatin
Prominent cytoplasmic structures are the
electron-dense granules = membrane-
bound and contain preformed mediators
Ultrastructure

Crystalline structures pattern of
membrane-bound secretory granules:
Scrolls

Ultrastructure

Grating

Ultrastructure

Lattices

Appearance in tissue
Images from: http://www.pathologyoutlines.com/topic/bonemarrowmastcells.html
Can be round or spindle-shaped

Most effective way to identify the
location and subtype histologically =
immunohistochemical analysis using Ab
against mast cellspecific proteases
Granule matrix

Formed from proteoglycans, with
glycosaminoglycan (GAG) side chains
Heparin = main proteoglycans in human
mast cells
Stabilizes the -tryptase tetramer
Neutral proteases, acid hydrolases, and
histamine molecules are attached to
heparin by ionic linkage to the sulfate
groups on the GAGs.
Some chondroitin E also present
Preform mediators

Release of mediators

Degranulation from activation
Energy-dependent
After almost complete degranulation,
HLMCs are able to survive and undergo
regranulation over a period of 48 hours

Piecemeal degranulation
Poorly understood mechanism
Variable loss of granule contents
Granules and their membranes remain
intact


Synthesis of new mediators


Synthesis and secretion of cytokines

Synthesis and secretion of cytokines

Synthesis and secretion of chemokines
CCL1-7
CCL12
CCL17
CCL19
CCL20
CCL22
CXCL5
CXCL8

Image from: http://www.biochemj.org/csb/011/Fig11_mast_cell_signallinga.jpg

F I Hsu, J A Boyce, Middletons Allergy 7th edition, 2009, 311-328.
Activation mechanisms
IgE-dependent activation
Monomeric IgE-dependent
activation
Non-immunologic mast cell
activation
Activation mechanisms

Toshiaki Kawakami & Stephen J. Galli, Nature Reviews Immunology 2, 2002, 773-786
Initiate through the high-affinity IgE
receptor FcRI

Stephen J. Galli, Mindy Tsai & Adrian M. Piliponsky, Nature 454 (2008), 445-454
Granule swelling
Crystal dissolution
Granule fusion with surrounding
granules and cell membrane
Exocytosis + release of mediators
into the extracellular space

Microscopic process

Inhibition measures
Mast cell stablilizer
Target: 1 of LPA receptors, GPR35
Poor efficacy in vivo
Rapid tachyphylaxis
2-adrenoceptor agonists
Poor efficacy in vivo
Rapid tachyphylaxis
Syk inhibitor
Poor outcome
Monomeric IgE activation

Binding of monomeric IgE alone to FcRI
initiates intracellular signaling events
and Ca2+ influx
In HCBMCs, monomeric IgE binding
induces the release of CCL1, CCL3, and
GM-CSF without histamine release
In HLMCs, IgE binding induces secretion
of histamine, LTC4, and CXCL8, which is
markedly enhanced in the presence of
SCF
Monomeric IgE activation

In HLMCs
Ongoing signaling is dependent on the
presence of free IgE
Signaling ceases immediately when free IgE is
removed
SCF and free IgE concentrations are
elevated in asthmatic airways
Good correlation has been found
between total serum IgE and presence of
asthma and bronchial
hyperresponsiveness

Non-immunologic activation


SCF inhibits 2-adrenoceptor (2-
AR) signaling in HLMCs and HMC-1
within minutes of exposure,
followed by internalization.

=> Impaired 2-ARdependent
inhibition of
Histamine and LTC
4
release
Ion channel modulation

Activation via TLR-2 induces Ca2+
mobilization, degranulation and
LTC4 production

Activation via TLR3 can
deteriorate airway physiology

From mouse model and in vitro study,
mast cells could contribute to Th2
differentiation at the onset of an immune
response
Bee venom phospholipase (PL)A
2
and Der p 1
induce the release of histamine and IL-4 from
HLMCs in the absence of cell-bound IgE
Cockroaches, fungal spores, pollens, and cats
can induce the release of phospholipases and
proteases


Why allergen sensitization does not occur
in everyone?
Environmental factors
Level of allergen exposure
Genetic factors
Mast cell releasability
Epithelial integrity and permeability
Local antiprotease activity
Regulation of cytokine production

Influence on development of dendritic
cells and their ability to activate T cells
Histamine and PGD2 increases IL-10 and
decreases IL-12 production by mature
dendritic cells -> naive T cells become
polarized toward Th2 phenotype
Mast cell dependence for the generation of
Th2-promoting dendritic cells is evident in
mice

Influence on development of dendritic
cells and their ability to activate T cells
Mast cell exosomes induce immature
dendritic cells to become mature
plasmacytoid dendritic cells capable of
antigen presentation by upregulating MHC
class II, CD80, CD86, and CD40 molecules
Mast cellderived TNF- is important for
dendritic cell migration during immune
responses.
Roles in allergic diseases:
Anaphylaxis
Roles in anaphylaxis

Anaphylaxis is mediated predominantly
by mast cells tryptase
-tryptase
Released by mast cells constitutively
Increased baseline release in mastocytosis
-tryptase
Stored in mast cell granules
Released after IgE-dependent activation
> More specific marker than total tryptase.
> BEST marker of systemic mast cell activation in
anaphylaxis

Roles in anaphylaxis

Why does systemic activation of mast cell
occurred?
Systemic diffusion of allergen? Unlikely
Amplification mechanisms?
Neurologic reflexes
Platelet-activating factor (PAF)
Can activate human mast cells
Can cause mast cells to release histamine
Induces the release of CXCL8
Transiently upregulates mRNA expression for several
other chemokines
Enhances IgE-dependent mediator release

Allergic rhinitis
Roles in AR

No. of mast cells in the epithelium
Expression of Th2 cytokines in mast
cells
No. of CD34+, tryptase-negative cells
(mast cell progenitor) in the nasal
epithelium
Expression of IL-4, which is reversed by
the application of topical corticosteroids
Roles in AR

AR = IgE-driven, mast celldependent
disease
Histamine are not elevated, but
antihistamine therapy is highly effective at
ameliorating symptoms
Anti-IgE therapy also is effective
Ongoing mast cell activation in nasal mucosa
+ Biologic effects of mast cell products can
explain much of the symptomatology and
pathology of AR
Allergic conjunctivitis
Roles in AC

No. of mast cells and evidence of
degranulation in all types of AC
Levels of histamine, tryptase, and LTC4
are found in tears after allergen exposure
No. Of MC
T
cell found in conjunctival
epithelium and subepithelial layers of
PAC, SAC, and VKC patient
No. Of MC
TC
cells AKC and ABC patient
Atopic dermatitis and urticaria
Roles in atopic dermatitis

No. of MC
T
increases in the skin of
patients with atopic dermatitis
Expression of IL-4 of skin mast cells in
atopic dermatitis patient
Roles in urticaria

In acute urticaria
Mast cell degranulation is evident
Antihistamines is useful treatment,
suggesting that the skin lesions result from
mast cell activation
In CIU, mast cell activation is a factor
Constitutive histamine release compared
with control subjects
30% of patients have autoAb to FcRI or IgE
Anti-IgE (omalizumab) is highly effective
treatment
Asthma
Experimentally induced asthma

Late asthmatic reaction (LAR):
4 - 12 hr
Early asthmatic reaction (EAR):
10 min 2 hr
Bronchial allergen challenge
Then, check the fall of FEV1
Early asthmatic reaction

Mediator release from HLMCs in vitro:

Half-maximal release occurring

Similar pattern found in bronchoalveolar
lavage
Histamine PGD2 LTC4
2 min 5 min 10 min

Histamine, PGD2, and LTC4/LTD4 induce
bronchoconstriction, mucosal edema, and
mucus secretion

EAR was markedly attenuated by
inhibitors of
Histamine (H1 receptor)
LTC4/LTD4 (cysteinyl LTRl)
To a lesser extent, PGD2 (thromboxane TP
receptor).


Evidences supporting mast cell origin:
Kinetics of IgE-dependent mediator release
in vivo parallels that of HLMC in vitro
Rapidly increased concentration of mast cell
specific tryptase in BAL occurs after local
bronchial allergen challenge
-agonists, when applied acutely in vitro,
completely abolish EAR and associated
increase in plasma histamine levels
EAR is almost completely ablated after 12 to
16 weeks of pretreatment with omalizumab,
Late asthmatic reaction

Associated with inflammatory cell
accumulation and activation
Concentrations of histamine, PGD2, and
LTC4
But in different ratios than during the EAR
Tryptase levels fall
GM-CSF
Released after allergen provocation
Inhibits expression of tryptase in HMC-1 cells but
does not attenuate histamine release
IgE-dependent histamine release in HLMCs
LAR is attenuated markedly by omalizumab
Role in chronic allergic asthma

Mast cells present in the bronchial
mucosal are in an activated state
Degranulation is continuous
No.of mast cells in BAL fluid
Histamine and tryptase
Expression of IL-4 and IL-5 mRNA in
mast cells
Expression of mast cellassociated IL-4
and TNF-
Role in chronic allergic asthma

Enhanced IgE-dependent release
Higher IgE concentrations
Upregulation of FcRI
Enhanced IgE-related signaling
Enhanced allergen-dependent mediator
release
In conclusion:
Atopic asthmatic phenotype = interaction
among allergens, IgE, and hyperreactive mast
cells
Role in non-allergic asthma

Mast cell FcRI+ expression in
bronchial mucosa, may be due to
Epsilon germline gene (I) and mature
epsilon heavy chain (C) mRNA+ B cells in the
bronchial mucosa
So local IgE synthesis
Expression of Th2 cytokines IL-4 and IL-
5 occurs at both mRNA and protein levels
Accordingly, anti-IgE therapy may
potentially be very effective, too
Role in occupational asthma

Pathology of occupational asthma (with
the exception of irritant-induced asthma)
is virtually identical to that seen in atopic
and intrinsic asthma
Role in exercise-induced asthma

This is not a distinct disease entity, but a
marker of poor asthma control and
ongoing airway inflammation

Agents that might help:
Histamine H1 receptor antagonists
Cyclooxygenase (COX) inhibitors
LTRA
Cromolyn sodium
Role in ASA-induced asthma

It is associated with LTC4 in nasal
secretions and LTE4 in urine

Mast cell LT generation may be involved
No.of mast cells in the airways
Proportion of these mast cells express
COX-2
Mast cells are the predominant cells
expressing LTC4 synthase
Role in asthma exacerbations

RSV can induce mast cell degranulation
Evidence that mast cells contribute
directly to asthma exacerbations
Activation of mast cells via TLR3, induces
secretion of both IFN- and IFN- ,
Dual stimulation through TLR3 and FcRI
enhances the release of IL-1, TNF-, IL-5,
and cysteinyl leukotrienes
Omalizumab significantly reduces the rate of
severe exacerbations
Mast cell location in asthma

No.of mast cells in the lamina propria is
not increased in asthmatic airway

But in asthmatic patient, mast cells
infiltrate 3 key structures
Airway epithelium
Airway mucosal glands
Airway smooth muscle (ASM)
Interaction with ASM

Interaction with epithelium

Interaction with fibroblasts

Mast cells have the potential to activate
subepithelial myofibroblasts
Mast cells and fibroblasts interact
intimately through several mechanisms
Histamine, basic FGF, and IL-4 promote
fibroblast proliferation in humans
IL-4 is a chemoattractant for human
fibroblasts and also induces fibroblasts to
secrete collagen type I, III and fibronectin
Interaction with fibroblasts

IL-13 increases CCL11 release from
human airway fibroblasts
Heparin stabilizes basic FGF structurally
and preserves its bioactivity by protecting
it from degradation
Thereby potentiating fibroblast activation
and proliferation indirectly

Take Home Messages
Mast cells are tissue-resident immune
cells, with wide array of function in
response to various stimuli
They are capable to secrete numerous
multifunctional substances (autocoid,
protease, cytokines, chemokines)
They play important roles in host
defense and in allergic diseases
They have complex interactions with
other immunologic and structural cells
Thank you

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